Regulation of CD8+ T cells undergoing primary and secondary responses to infection in the same host

Naive Ag-specific CD8(+) T cells expand, contract, and become memory cells after infection and/or vaccination. Memory CD8(+) T cells provide faster, more effective secondary responses against repeated exposure to the same pathogen. Using an adoptive transfer system with low numbers of trackable nontransgenic memory CD8(+) T cells, we showed that secondary responses can be comprised of both primary (naive) and secondary (memory) CD8(+) T cells after bacterial (Listeria monocytogenes) and/or viral (lymphocytic choriomeningitis virus) infections. The level of memory CD8(+) T cells present at the time of infection inversely correlated with the magnitude of primary CD8(+) T cell responses against the same epitope but directly correlated with the level of protection against infection. However, similar numbers of Ag-specific CD8(+) T cells were found 8 days postinfection no matter how many memory cells were present at the time of infection. Rapid contraction of primary CD8(+) T cell responses was not influenced by the presence of memory CD8(+) T cells. However, contraction of secondary CD8(+) T cell responses was markedly prolonged compared with primary responses in the same host mice. This situation occurred in response to lymphocytic choriomeningitis virus or L. monocytogenes infection and for CD8(+) T cell responses against multiple epitopes. The delayed contraction of secondary CD8(+) T cells was also observed after immunization with peptide-coated dendritic cells. Together, the results show that the level of memory CD8(+) T cells influences protective immunity and activation of naive precursors specific for the same epitope but has little impact on the magnitude or program of the CD8(+) T cell response.     

Cellular and humoral immunity against vaccinia virus infection of mice

Despite the widespread use of vaccinia virus (VV) as a vector for other Ags and as the smallpox vaccine, there is little information available about the protective components of the immune response following VV infection. In this study, protection against wild-type VV was evaluated in mice with respect to the relative contributions of CD8(+) T cells vs that of CD4(+) T cells and Ab. C57BL/6 mice primed with the Western Reserve strain of VV mount significant IgM and IgG Ab responses, specific cytotoxic T cell responses, IFN-gamma responses in CD4(+) and CD8(+) T cells, and effectively clear the virus. This protection was abrogated by in vivo depletion of CD4(+) T cells or B cells in IgH(-/-) mice, but was not sensitive to CD8(+) T cell depletion alone. However, a role for CD8(+) T cells in primary protection was demonstrated in MHC class II(-/-) mice, where depleting CD8(+) T cells lead to increase severity of disease. Unlike control MHC class II(-/-) mice, the group depleted of CD8(+) T cells developed skin lesions on the tail and feet and had adrenal necrosis. Adoptive transfer experiments also show CD8(+) T cells can mediate protective memory. These results collectively show that both CD4(+) and CD8(+) T cell-mediated immunity can contribute to protection against VV infection. However, CD4(+) T cell-dependent anti-virus Ab production plays a more important role in clearing virus following acute infection, while in the absence of Ab, CD8(+) T cells can contribute to protection against disease.     

IL-17A-producing gammadeltaT cells promote CTL responses against Listeria monocytogenes infection by enhancing dendritic cell cross-presentation

Interleukin-17A-producing T cells, especially Th17, have been shown to be involved in inflammatory autoimmune diseases and host defense against extracellular infections. However, whether and how IL-17A or IL-17A-producing cells can help protection against intracellular bacteria remains controversial, especially how it regulates the adaptive immunity besides recruitment of neutrophils in the innate immune system. By infecting IL-17A-deficient mice with Listeria monocytogenes, we show in this study that IL-17A is required for the generation of Ag-specific CD8(+) CTL response against primary infection, but not for the generation of memory CD8(+) T cells against secondary challenge. Interestingly, we identify γδT cells, but not conventional CD4(+) Th17 cells, as the main cells for innate IL-17A production during L. monocytogenes infection. Furthermore, γδT cells are found to promote Ag-specific CD8(+) T cell proliferation by enhancing cross-presentation of dendritic cells through IL-17A. Adoptive transfer of Il17a(+/+) γδT cells, but not Il17a(-/-) γδT cells or Il17a(+/+) CD4(+) T cells, were sufficient to recover dendritic cells cross-presentation and defective CD8(+) T cell response in Il17a(-/-) mice. Our findings indicate an important role of infection-inducible IL-17A-producing γδT cells and their derived IL-17A against intracellular bacterial infection, providing a mechanism of IL-17A for regulation of innate and adaptive immunity.     

Persistent infection with ebola virus under conditions of partial immunity

Ebola hemorrhagic fever in humans is associated with high mortality; however, some infected hosts clear the virus and recover. The mechanisms by which this occurs and the correlates of protective immunity are not well defined. Using a mouse model, we determined the role of the immune system in clearance of and protection against Ebola virus. All CD8 T-cell-deficient mice succumbed to subcutaneous infection and had high viral antigen titers in tissues, whereas mice deficient in B cells or CD4 T cells cleared infection and survived, suggesting that CD8 T cells, independent of CD4 T cells and antibodies, are critical to protection against subcutaneous Ebola virus infection. B-cell-deficient mice that survived the primary subcutaneous infection (vaccinated mice) transiently depleted or not depleted of CD4 T cells also survived lethal intraperitoneal rechallenge for >/==" BORDER="0">25 days. However, all vaccinated B-cell-deficient mice depleted of CD8 T cells had high viral antigen titers in tissues following intraperitoneal rechallenge and died within 6 days, suggesting that memory CD8 T cells by themselves can protect mice from early death. Surprisingly, vaccinated B-cell-deficient mice, after initially clearing the infection, were found to have viral antigens in tissues later (day 120 to 150 post-intraperitoneal infection). Furthermore, following intraperitoneal rechallenge, vaccinated B-cell-deficient mice that were transiently depleted of CD4 T cells had high levels of viral antigen in tissues earlier (days 50 to 70) than vaccinated undepleted mice. This demonstrates that under certain immunodeficiency conditions, Ebola virus can persist and that loss of primed CD4 T cells accelerates the course of persistent infections. These data show that CD8 T cells play an important role in protection against acute disease, while both CD4 T cells and antibodies are required for long-term protection, and they provide evidence of persistent infection by Ebola virus suggesting that under certain conditions of immunodeficiency a host can harbor virus for prolonged periods, potentially acting as a reservoir.     

Multiple Chlamydia pneumoniae antigens prime CD8+ Tc1 responses that inhibit intracellular growth of this vacuolar pathogen

CD8(+) T cells play an essential role in immunity to Chlamydia pneumoniae (Cpn). However, the target Ags recognized by Cpn-specific CD8(+) T cells have not been identified, and the mechanisms by which this T cell subset contributes to protection remain unknown. In this work we demonstrate that Cpn infection primes a pathogen-specific CD8(+) T cell response in mice. Eighteen H-2(b) binding peptides representing sequences from 12 Cpn Ags sensitized target cells for MHC class I-restricted lysis by CD8(+) CTL generated from the spleens and lungs of infected mice. Peptide-specific IFN-gamma-secreting CD8(+) T cells were present in local and systemic compartments after primary infection, and these cells expanded after pathogen re-exposure. CD8(+) T cell lines to the 18 Cpn epitope-bearing peptides were cytotoxic, displayed a memory phenotype, and secreted IFN-gamma and TNF-alpha, but not IL-4. These CTL lines lysed Cpn-infected macrophages, and the lytic activity was inhibited by brefeldin A, indicating endogenous processing of CTL Ags. Finally, Cpn peptide-specific CD8(+) CTL suppressed chlamydial growth in vitro by direct lysis of infected cells and by secretion of IFN-gamma and other soluble factors. These studies provide information on the mechanisms by which CD8(+) CTL protect against Cpn, furnish the tools to investigate their possible role in immunopathology, and lay the foundation for future work to develop vaccines against acute and chronic Cpn infections.     

CD4+ T-cell responses are required for clearance of West Nile virus from the central nervous system

Although studies have established that innate and adaptive immune responses are important in controlling West Nile virus (WNV) infection, the function of CD4(+) T lymphocytes in modulating viral pathogenesis is less well characterized. Using a mouse model, we examined the role of CD4(+) T cells in coordinating protection against WNV infection. A genetic or acquired deficiency of CD4(+) T cells resulted in a protracted WNV infection in the central nervous system (CNS) that culminated in uniform lethality by 50 days after infection. Mice surviving past day 10 had high-level persistent WNV infection in the CNS compared to wild-type mice, even 45 days following infection. The absence of CD4(+) T-cell help did not affect the kinetics of WNV infection in the spleen and serum, suggesting a role for CD4-independent clearance mechanisms in peripheral tissues. WNV-specific immunoglobulin M (IgM) levels were similar to those of wild-type mice in CD4-deficient mice early during infection but dropped approximately 20-fold at day 15 postinfection, whereas IgG levels in CD4-deficient mice were approximately 100- to 1,000-fold lower than in wild-type mice throughout the course of infection. WNV-specific CD8(+) T-cell activation and trafficking to the CNS were unaffected by the absence of CD4(+) T cells at day 9 postinfection but were markedly compromised at day 15. Our experiments suggest that the dominant protective role of CD4(+) T cells during primary WNV infection is to provide help for antibody responses and sustain WNV-specific CD8(+) T-cell responses in the CNS that enable viral clearance.     

Protection from bacterial infection by a single vaccination with replication-deficient mutant herpes simplex virus type 1

Adaptive immune responses in which CD8(+) T cells recognize pathogen-derived peptides in the context of major histocompatibility complex class I molecules play a major role in the host defense against infection with intracellular pathogens. Cells infected with intracellular bacteria such as Listeria monocytogenes, Salmonella enterica serovar Typhimurium, or Mycobacterium tuberculosis are directly lysed by cytotoxic CD8(+) T cells. For this reason, current vaccines for intracellular pathogens, such as subunit vaccines or viable bacterial vaccines, aim to generate robust cytotoxic T-cell responses. In order to investigate the capacity of a herpes simplex virus type 1 (HSV-1) vector to induce strong cytotoxic effector cell responses and protection from infection with intracellular pathogens, we developed a replication-deficient, recombinant HSV-1 (rHSV-1) vaccine. We demonstrate in side-by-side comparison with DNA vaccination that rHSV-1 vaccination induces very strong CD8(+) effector T-cell responses. While both vaccines provided protection from infection with L. monocytogenes at low, but lethal doses, only rHSV-1 vaccines could protect from higher infectious doses; HSV-1 induced potent memory cytotoxic T lymphocytes that, upon challenge by pathogens, efficiently protected the animals. Despite the stimulation of relatively low humoral and CD4-T-cell responses, rHSV-1 vectors are strong candidates for future vaccine strategies that confer efficient protection from subsequent infection with intracellular bacteria.     

Generation of a convalescent model of virulent Francisella tularensis infection for assessment of host requirements for survival of tularemia

Francisella tularensis is a facultative intracellular bacterium and the causative agent of tularemia. Development of novel vaccines and therapeutics for tularemia has been hampered by the lack of understanding of which immune components are required to survive infection. Defining these requirements for protection against virulent F. tularensis, such as strain SchuS4, has been difficult since experimentally infected animals typically die within 5 days after exposure to as few as 10 bacteria. Such a short mean time to death typically precludes development, and therefore assessment, of immune responses directed against virulent F. tularensis. To enable identification of the components of the immune system that are required for survival of virulent F. tularensis, we developed a convalescent model of tularemia in C57Bl/6 mice using low dose antibiotic therapy in which the host immune response is ultimately responsible for clearance of the bacterium. Using this model we demonstrate αβTCR(+) cells, γδTCR(+) cells, and B cells are necessary to survive primary SchuS4 infection. Analysis of mice deficient in specific soluble mediators shows that IL-12p40 and IL-12p35 are essential for survival of SchuS4 infection. We also show that IFN-γ is required for survival of SchuS4 infection since mice lacking IFN-γR succumb to disease during the course of antibiotic therapy. Finally, we found that both CD4(+) and CD8(+) cells are the primary producers of IFN-γand that γδTCR(+) cells and NK cells make a minimal contribution toward production of this cytokine throughout infection. Together these data provide a novel model that identifies key cells and cytokines required for survival or exacerbation of infection with virulent F. tularensis and provides evidence that this model will be a useful tool for better understanding the dynamics of tularemia infection.     

Long-term control of Mycobacterium tuberculosis infection is mediated by dynamic immune responses

The primary goal of this study was to determine how chronic exposure to Ag influences the functionality of Mycobacterium tuberculosis-specific T cell responses. The frequency of IFN-gamma-producing effector CD4(+) and CD8(+) T cells dynamically changed during persistent M. tuberculosis infection. CD8(+) T cells used differential effector functions during acute and chronic phases of the immune response, where CD8(+) T cells produced negligible amounts of IFN-gamma early in infection, but switched to cytokine production during the chronic stage of infection. Using limiting dilution analysis, CD8(+) T cells isolated during the initial phase of infection demonstrated lytic potential, but this waned in the chronic stage. The apparent loss of cytotoxic activity was not associated with the lack of perforin. Ag dose could potentially govern the functional program of CD8(+) T cells. Collectively, these results depict a host immune response mounted against M. tuberculosis of a significantly more dynamic nature than previously recognized.     

Expansion of protective CD8+ T-cell responses driven by recombinant cytomegaloviruses

CD8(+) T cells are critical for the control of many persistent viral infections, such as human immunodeficiency virus, hepatitis C virus, Epstein-Barr virus, and cytomegalovirus (CMV). In most infections, large CD8(+)-T-cell populations are induced early but then contract and are maintained thereafter at lower levels. In contrast, CD8(+) T cells specific for murine CMV (MCMV) have been shown to gradually accumulate after resolution of primary infection. This unique behavior is restricted to certain epitopes, including an immunodominant epitope derived from the immediate-early 1 (IE1) gene product. To explore the mechanism behind this further, we measured CD8(+)-T-cell-mediated immunity induced by recombinant MCMV-expressing epitopes derived from influenza A virus or lymphocytic choriomeningitis virus placed under the control of an IE promoter. We observed that virus-specific CD8(+)-T-cell populations were induced and that these expanded gradually over time. Importantly, these CD8(+) T cells provided long-term protection against challenge without boosting. These results demonstrate a unique pattern of accumulating T cells, which provide long-lasting immune protection, that is independent of the initial immunodominance of the epitope and indicates the potential of T-cell-inducing vaccines based on persistent vectors.     

Vaccination against lymphocytic choriomeningitis virus infection in MHC class II-deficient mice

The impact of prophylactic vaccination against acute and chronic infection in a Th-deficient host has not been adequately addressed because of difficulties in generating protective immunity in the absence of CD4(+) T cell help. In this study, we demonstrated that a broad CD8(+) T cell immune response could be elicited in MHC class II-deficient mice by vaccination with adenovirus encoding lymphocytic choriomeningitis virus (LCMV) glycoprotein tethered to MHC class II-associated invariant chain. Moreover, the response induced conferred significant cytolytic CD8(+) T cell-mediated protection against challenge with a high dose of the invasive clone 13 strain of LCMV. In contrast, vaccination with adenovirus encoding unlinked LCMV glycoprotein induced weak virus control in the absence of CD4(+) T cells, and mice may die of increased immunopathology associated with incomplete protection. Acute mortality was not observed in any vaccinated mice following infection with the less-invasive Traub strain. However, LCMV Traub infection caused accelerated late mortality in unvaccinated MHC class II-deficient mice; in this case, we observed a strong trend toward delayed mortality in vaccinated mice, irrespective of the nature of the vaccine. These results indicated that optimized vaccination may lead to efficient protection against acute viral infection, even in Th-deficient individuals, but that the duration of such immunity is limited. Nevertheless, for select immunodeficiencies in which CD4(+) T cell deficiency is incomplete or transient, these results are very encouraging.     

Late priming and variability of epitope-specific CD8+ T cell responses during a persistent virus infection

Control of persistently infecting viruses requires that antiviral CD8(+) T cells sustain their numbers and effector function. In this study, we monitored epitope-specific CD8(+) T cells during acute and persistent phases of infection by polyoma virus, a mouse pathogen that is capable of potent oncogenicity. We identified several novel polyoma-specific CD8(+) T cell epitopes in C57BL/6 mice, a mouse strain highly resistant to polyoma virus-induced tumors. Each of these epitopes is derived from the viral T proteins, nonstructural proteins produced by both productively and nonproductively (and potentially transformed) infected cells. In contrast to CD8(+) T cell responses described in other microbial infection mouse models, we found substantial variability between epitope-specific CD8(+) T cell responses in their kinetics of expansion and contraction during acute infection, maintenance during persistent infection, as well as their expression of cytokine receptors and cytokine profiles. This epitope-dependent variability also extended to differences in maturation of functional avidity from acute to persistent infection, despite a narrowing in TCR repertoire across all three specificities. Using a novel minimal myeloablation-bone marrow chimera approach, we visualized priming of epitope-specific CD8(+) T cells during persistent virus infection. Interestingly, epitope-specific CD8(+) T cells differed in CD62L-selectin expression profiles when primed in acute or persistent phases of infection, indicating that the context of priming affects CD8(+) T cell heterogeneity. In summary, persistent polyoma virus infection both quantitatively and qualitatively shapes the antiviral CD8(+) T cell response.     

Macrophages, CD4+ or CD8+ cells are each sufficient for protection against Chlamydia pneumoniae infection through their ability to secrete IFN-gamma

By using a T, B, or NK cell-deficient mouse strain (recombinase-activating gene (RAG)-1(-/-)/common cytokine receptor gamma-chain (gamma(C)R)), and T and B cell and IFN-gamma-deficient (RAG-1(-/-)/IFN-gamma(-/-)) mice, we have studied the generation of immunity against infection by Chlamydia pneumoniae. We found that IFN-gamma secreted by innate-cell populations protect against C. pneumoniae infection. However, NK cells were not needed for such IFN-gamma-dependent innate immune protection. Inoculation of wild type, but not IFN-gamma(-/-) bone marrow-derived macrophages protected RAG-1(-/-)/IFN-gamma(-/-) mice against C. pneumoniae infection. In line, pulmonary macrophages from RAG-1(-/-) C. pneumoniae-infected mice expressed IFN-gamma mRNA. Reconstitution of RAG-1(-/-)/gamma(c)R(-/-) or RAG-1(-/-)/IFN-gamma(-/-) mice with CD4(+) or CD8(+) cells by i.v. transfer of FACS sorted wild type spleen cells (SC) increased resistance to C. pneumoniae infection. On the contrary, no protection was observed upon transfer of IFN-gamma(-/-) CD4(+) or IFN-gamma(-/-) CD8(+) SC. T cell-dependent protection against C. pneumoniae was weaker when IFN-gammaR(-/-) CD4(+) or IFN-gammaR(-/-) CD8(+) SC were inoculated into RAG-1(-/-)/IFN-gamma(-/-) mice. Thus both nonlymphoid and T cell-derived IFN-gamma can play a central and complementary role in protection against C. pneumoniae. IFN-gamma secreted by nonlymphoid cells was not required for T cell-mediated protection against C. pneumoniae; however, IFN-gamma regulated T cell protective functions.     

Foxp3+ regulatory T cells control persistence of viral CNS infection

We earlier established a model of a persistent viral CNS infection using two week old immunologically normal (genetically unmodified) mice and recombinant measles virus (MV). Using this model infection we investigated the role of regulatory T cells (Tregs) as regulators of the immune response in the brain, and assessed whether the persistent CNS infection can be modulated by manipulation of Tregs in the periphery. CD4(+) CD25(+) Foxp3(+) Tregs were expanded or depleted during the persistent phase of the CNS infection, and the consequences for the virus-specific immune response and the extent of persistent infection were analyzed. Virus-specific CD8(+) T cells predominantly recognising the H-2D(b)-presented viral hemagglutinin epitope MV-H(22-30) (RIVINREHL) were quantified in the brain by pentamer staining. Expansion of Tregs after intraperitoneal (i.p.) application of the superagonistic anti-CD28 antibody D665 inducing transient immunosuppression caused increased virus replication and spread in the CNS. In contrast, depletion of Tregs using diphtheria toxin (DT) in DEREG (depletion of regulatory T cells)-mice induced an increase of virus-specific CD8(+) effector T cells in the brain and caused a reduction of the persistent infection. These data indicate that manipulation of Tregs in the periphery can be utilized to regulate virus persistence in the CNS.     

Stable CD8+ T cell memory during persistent Trypanosoma cruzi infection

CD8(+) T cell responses to persistent infections caused by intracellular pathogens are dominated by resting T effectors and T effector memory cells, with little evidence suggesting that a T central memory (T(CM)) population is generated. Using a model of Trypanosoma cruzi infection, we demonstrate that in contrast to the T effector/T effector memory phenotype of the majority of T. cruzi-specific CD8(+) T cells, a population of cells displaying hallmark characteristics of T(CM) cells is also present during long-term persistent infection. This population expressed the T(CM) marker CD127 and a subset expressed one or more of three other T(CM) markers: CD62L, CCR7, and CD122. Additionally, the majority of CD127(high) cells were KLRG1(low), indicating that they have not been repetitively activated through TCR stimulation. These CD127(high) cells were better maintained than their CD127(low) counterparts following transfer into naive mice, consistent with their observed surface expression of CD127 and CD122, which confer the ability to self-renew in response to IL-7 and IL-15. CD127(high) cells were capable of IFN-gamma production upon peptide restimulation and expanded in response to challenge infection, indicating that these cells are functionally responsive upon Ag re-encounter. These results are in contrast to what is typically observed during many persistent infections and indicate that a stable population of parasite-specific CD8(+) T cells capable of Ag-independent survival is maintained in mice despite the presence of persistent Ag.     

CD8+ cellular immunity mediates rAd5 vaccine protection against Ebola virus infection of nonhuman primates

Vaccine-induced immunity to Ebola virus infection in nonhuman primates (NHPs) is marked by potent antigen-specific cellular and humoral immune responses; however, the immune mechanism of protection remains unknown. Here we define the immune basis of protection conferred by a highly protective recombinant adenovirus virus serotype 5 (rAd5) encoding Ebola virus glycoprotein (GP) in NHPs. Passive transfer of high-titer polyclonal antibodies from vaccinated Ebola virus-immune cynomolgus macaques to naive macaques failed to confer protection against disease, suggesting a limited role of humoral immunity. In contrast, depletion of CD3(+) T cells in vivo after vaccination and immediately before challenge eliminated immunity in two vaccinated macaques, indicating a crucial requirement for T cells in this setting. The protective effect was mediated largely by CD8(+) cells, as depletion of CD8(+) cells in vivo using the cM-T807 monoclonal antibody (mAb), which does not affect CD4(+) T cell or humoral immune responses, abrogated protection in four out of five subjects. These findings indicate that CD8(+) cells have a major role in rAd5-GP-induced immune protection against Ebola virus infection in NHPs. Understanding the immunologic mechanism of Ebola virus protection will facilitate the development of vaccines for Ebola and related hemorrhagic fever viruses in humans.     

Listeria monocytogenes infection overcomes the requirement for CD40 ligand in exogenous antigen presentation to CD8(+) T cells.

In vivo priming of CD8(+) T lymphocytes against exogenously processed model Ags requires CD4(+) T cell help, specifically interactions between CD40 ligand (CD40L) expressed by activated CD4(+) T cells and CD40, which is present on professional APC such as dendritic cells (DCs). To address this issue in the context of bacterial infection, we examined CD40L-CD40 interactions in CD8(+) T cell priming against an exogenously processed, nonsecreted bacterial Ag. CD40L interactions were blocked by in vivo treatment with anti-CD40L mAb MR-1, which inhibited germinal center formation and CD8(+) T cell cross-priming against an exogenous model Ag, OVA. In contrast, MR-1 treatment did not interfere with CD8(+) T cell priming against a nonsecreted or secreted recombinant Ag expressed by Listeria monocytogenes. Memory and secondary responses of CD8(+) T cells against nonsecreted and secreted bacterial Ags were also largely unimpaired by transient MR-1 treatment. When MR-1-treated mice were concurrently immunized with L. monocytogenes and OVA-loaded splenocytes, cross-priming of OVA-specific naive CD8(+) T cells occurred. No significant decline in cross-priming against OVA was measured when either TNF or IFN-gamma was neutralized in L. monocytogenes-infected animals, demonstrating that multiple signals exist to overcome CD40L blockade of CD8(+) T cell cross-priming during bacterial infection. These data support a model in which DCs can be stimulated in vivo through signals other than CD40, becoming APC that can effectively stimulate CD8(+) T cell responses against exogenous Ags during infection.     

The antiviral CD8+ T cell response is differentially dependent on CD4+ T cell help over the course of persistent infection

Although many studies have investigated the requirement for CD4(+) T cell help for CD8(+) T cell responses to acute viral infections that are fully resolved, less is known about the role of CD4(+) T cells in maintaining ongoing CD8(+) T cell responses to persistently infecting viruses. Using mouse polyoma virus (PyV), we asked whether CD4(+) T cell help is required to maintain antiviral CD8(+) T cell and humoral responses during acute and persistent phases of infection. Though fully intact during acute infection, the PyV-specific CD8(+) T cell response declined numerically during persistent infection in MHC class II-deficient mice, leaving a small antiviral CD8(+) T cell population that was maintained long term. These unhelped PyV-specific CD8(+) T cells were functionally unimpaired; they retained the potential for robust expansion and cytokine production in response to Ag rechallenge. In addition, although a strong antiviral IgG response was initially elicited by MHC class II-deficient mice, these Ab titers fell, and long-lived PyV-specific Ab-secreting cells were not detected in the bone marrow. Finally, using a minimally myeloablative mixed bone marrow chimerism approach, we demonstrate that recruitment and/or maintenance of new virus-specific CD8(+) T cells during persistent infection is impaired in the absence of MHC class II-restricted T cells. In summary, these studies show that CD4(+) T cells differentially affect CD8(+) T cell responses over the course of a persistent virus infection.     

Generation of antifungal effector CD8+ T cells in the absence of CD4+ T cells during Cryptococcus neoformans infection

Immunity to the opportunistic fungus Cryptococcus neoformans is dependent on cell-mediated immunity. Individuals with defects in cellular immunity, CD4(+) T cells in particular, are susceptible to infection with this pathogen. In host defense against a number of pathogens, CD8(+) T cell responses are dependent upon CD4(+) T cell help. The goal of these studies was to determine whether CD4(+) T cells are required for the generation of antifungal CD8(+) T cell effectors during pulmonary C. neoformans infection. Using a murine intratracheal infection model, our results demonstrated that CD4(+) T cells were not required for the expansion and trafficking of CD8(+) T cells to the site of infection. CD4(+) T cells were also not required for the generation of IFN-gamma-producing CD8(+) T cell effectors in the lungs. In CD4(-) mice, depletion of CD8(+) T cells resulted in increased intracellular infection of pulmonary macrophages by C. neoformans, increasing the pulmonary burden of the infection. Neutralization of IFN-gamma in CD4(-)CD8(+) mice similarly increased macrophage infection by C. neoformans, thereby blocking the protection provided by CD8(+) T cells. Altogether, these data support the hypothesis that effector CD8(+) T cell function is independent of CD4(+) T cells and that IFN-gamma production from CD8(+) T cells plays a role in controlling C. neoformans by limiting survival of C. neoformans within macrophages.     

Induction of humoral and CD8+ T cell responses are required for protection against lethal Ebola virus infection

Ebola virus (EBOV)-like particles (eVLP), composed of the EBOV glycoprotein and matrix viral protein (VP)40 with a lipid membrane, are a highly efficacious method of immunization against EBOV infection. The exact requirements for immunity against EBOV infection are poorly defined at this time. The goal of this work was to determine the requirements for EBOV immunity following eVLP vaccination. Vaccination of BALB/c or C57BL/6 mice with eVLPs in conjunction with QS-21 adjuvant resulted in mixed IgG subclass responses, a Th1-like memory cytokine response, and protection from lethal EBOV challenge. Further, this vaccination schedule led to the generation of both CD4(+) and CD8(+) IFN-gamma(+) T cells recognizing specific peptides within glycoprotein and VP40. The transfer of both serum and splenocytes, but not serum or splenocytes alone, from eVLP-vaccinated mice conferred protection against lethal EBOV infection in these studies. B cells were required for eVLP-mediated immunity to EBOV because B cell-deficient mice vaccinated with eVLPs were not protected from lethal EBOV challenge. We also found that CD8(+), but not CD4(+), T cells are absolutely required for eVLP-mediated protection against EBOV infection. Further, eVLP-induced protective mechanisms were perforin-independent, but IFN-gamma-dependent. Taken together, both EBOV-specific humoral and cytotoxic CD8(+) T cell responses are critical to mediate protection against filoviruses following eVLP vaccination.