Phenotypic and functional heterogeneity of EBV epitope-specific CD8+ T cells

High frequencies of EBV-specific CD8(+) T cells have been detected during acute EBV infection, yet persistent infection inevitably results. To address this issue, we characterized the phenotype and function of epitope-specific CD8(+) T cell populations from presentation with acute through latent infection. Considerable phenotypic and functional heterogeneity within, as well as between, two different epitope-specific populations was observed over time following acute infection. B7 EBV-encoded nuclear Ag (EBNA)-3A-specific CD8(+) T cells expressed only CD45RO from acute through latent EBV infection. A2 BMLF-1-specific CD8(+) T cells expressed CD45RO during acute infection and either CD45RA or CD45RO during latent EBV infection. This difference in CD45 isoform expression between the two epitope-specific populations did not translate into differences in perforin content, the ability to produce IFN-gamma, or the ability to proliferate in response to Ag in vitro. In individuals with latent EBV infection, the frequencies of A2 BMLF-1- or B7 EBNA-3A-specific CD8(+) T cells that expressed CD45RA, CD45RO, CD62 ligand, CCR7, and perforin were stable over time. However, the expression of CD62 ligand and CCR7 was significantly higher among EBNA-3A-specific CD8(+) T cells than among BMLF-1-specific CD8(+) T cells. Further work is necessary to understand how phenotypic and functional differences between EBV epitope-specific CD8(+) T cells are related to the biology of the virus and to the equilibrium between the virus and the host during persistent infection.     

Differential kinetics and specificity of EBV-specific CD4+ and CD8+ T cells during primary infection

The generation and maintenance of virus-specific CD4(+) T cells in humans are not well understood. We used short in vitro stimulation assays followed by intracellular cytokine staining to characterize the timing, magnitude, and Ag specificity of CD4(+) T cells over the course of primary EBV infection. Lytic and latent protein-specific CD4(+) T cells were readily detected at presentation with acute infectious mononucleosis and declined rapidly thereafter. Responses to BZLF-1, BMLF-1, and Epstein-Barr nuclear Ag-3A were more commonly detected than responses to Epstein-Barr nuclear Ag-1. Concurrent analyses of BZLF-1-specific CD4(+) and CD8(+) T cells revealed differences in the expansion, specificity, and stability of CD4(+) and CD8(+) T cell-mediated responses over time. Peripheral blood EBV load directly correlated with the frequency of EBV-specific CD4(+) T cell responses at presentation and over time, suggesting that EBV-specific CD4(+) T cell responses are Ag-driven.     

Increased frequency of EBV-specific effector memory CD8+ T cells correlates with higher viral load in rheumatoid arthritis

EBV is a candidate trigger of rheumatoid arthritis (RA). We determined both EBV-specific T cell and B cell responses and cell-associated EBV DNA copies in patients with RA and demographically matched healthy virus carriers. Patients with RA showed increased and broadened IgG responses to lytic and latent EBV-encoded Ags and 7-fold higher levels of EBV copy numbers in circulating blood cells. Additionally, patients with RA exhibited substantial expansions of CD8(+) T cells specific for pooled EBV Ags expressed during both B cell transformation and productive viral replication and the frequency of CD8(+) T cells specific for these Ags correlated with cellular EBV copy numbers. In contrast, CD4(+) T cell responses to EBV and T cell responses to human CMV Ags were unchanged, altogether arguing against a defective control of latent EBV infection in RA. Our data show that the regulation of EBV infection is perturbed in RA and suggest that increased EBV-specific effector T cell and Ab responses are driven by an elevated EBV load in RA.     

Differential evolution and stability of epitope-specific CD8(+) T cell responses in EBV infection

Murine models of lymphocytic choriomeningitis virus infection suggest that the memory CD8(+) T cell repertoire is reflective of the CD8(+) T cell repertoire generated during acute infection. Less is known regarding the evolution of CD8(+) T cell repertoires during human viral infections. We therefore examined epitope-specific CD8(+) T cell responses in a large cohort of individuals with acute through latent Epstein-Barr virus infection. Using 16 of 20 published EBV epitopes restricted by HLA-A2, HLA-A3 or HLA-B7, we showed that lytic cycle-specific CD8(+) T cell responses predominated during acute EBV infection. However, whereas HLA-A2(+)-restricted BMLF-1-specific CD8(+) T cell responses were maintained through latency, HLA-A2(+)- and HLA-B7(+)-restricted BZLF-1, as well as HLA-A3(+)-restricted BRLF-1 CD8(+) T cell responses, were generated but not readily maintained. Analyses of CD8(+) T cell responses to EBV latent cycle Ags showed delayed detection and lower frequencies of latent epitope-specific CD8(+) T cell responses during acute EBV infection, with maintenance of these responses 1 yr post-EBV infection. Early BMLF-1 and EBNA-3A epitope-specific CD8(+) T cell frequencies did not correlate with their frequencies at 1 yr postinfection. Interestingly, populations of EBV-specific CD8(+) T cells were stable during 20 mo in our long term EBV-seropositive populations, suggesting homeostasis between virus and the host immune system. This study demonstrates that CD8(+) T cell repertoires generated during persistent viral infections are not simply reflective of the initial pool of CD8(+) T cells and provides evidence that the generation of CD8(+) T cell responses to a persistent infection is a dynamic process.     

Increased CD8+ T Cell Response to Epstein-Barr Virus Lytic Antigens in the Active Phase of Multiple Sclerosis

It has long been known that multiple sclerosis (MS) is associated with an increased Epstein-Barr virus (EBV) seroprevalence and high immune reactivity to EBV and that infectious mononucleosis increases MS risk. This evidence led to postulate that EBV infection plays a role in MS etiopathogenesis, although the mechanisms are debated. This study was designed to assess the prevalence and magnitude of CD8+ T-cell responses to EBV latent (EBNA-3A, LMP-2A) and lytic (BZLF-1, BMLF-1) antigens in relapsing-remitting MS patients (n = 113) and healthy donors (HD) (n = 43) and to investigate whether the EBV-specific CD8+ T cell response correlates with disease activity, as defined by clinical evaluation and gadolinium-enhanced magnetic resonance imaging. Using HLA class I pentamers, lytic antigen-specific CD8+ T cell responses were detected in fewer untreated inactive MS patients than in active MS patients and HD while the frequency of CD8+ T cells specific for EBV lytic and latent antigens was higher in active and inactive MS patients, respectively. In contrast, the CD8+ T cell response to cytomegalovirus did not differ between HD and MS patients, irrespective of the disease phase. Marked differences in the prevalence of EBV-specific CD8+ T cell responses were observed in patients treated with interferon-β and natalizumab, two licensed drugs for relapsing-remitting MS. Longitudinal studies revealed expansion of CD8+ T cells specific for EBV lytic antigens during active disease in untreated MS patients but not in relapse-free, natalizumab-treated patients. Analysis of post-mortem MS brain samples showed expression of the EBV lytic protein BZLF-1 and interactions between cytotoxic CD8+ T cells and EBV lytically infected plasma cells in inflammatory white matter lesions and meninges. We therefore propose that inability to control EBV infection during inactive MS could set the stage for intracerebral viral reactivation and disease relapse.     

Epstein-Barr nuclear antigen 1-specific CD4(+) Th1 cells kill Burkitt's lymphoma cells

The gamma-herpesvirus, EBV, is reliably found in a latent state in endemic Burkitt's lymphoma. A single EBV gene product, Epstein-Barr nuclear Ag 1 (EBNA1), is expressed at the protein level. Several mechanisms prevent immune recognition of these tumor cells, including a block in EBNA1 presentation to CD8(+) killer T cells. Therefore, no EBV-specific immune response has yet been found to target Burkitt's lymphoma. We now find that EBNA1-specific, Th1 CD4(+) cytotoxic T cells recognize Burkitt's lymphoma lines. CD4(+) T cell epitopes of EBNA1 are predominantly found in the C-terminal, episome-binding domain of EBNA1, and approximately 0.5% of peripheral blood CD4(+) T cells are specific for EBNA1. Therefore, adaptive immunity can be directed against Burkitt's lymphoma, and perhaps this role for CD4(+) Th1 cells extends to other tumors that escape MHC class I presentation.     

Functional reversion of antigen-specific CD8+ T cells from patients with Hodgkin lymphoma following in vitro stimulation with recombinant polyepitope

Recent studies on Hodgkin's lymphoma (HL) have indicated that patients with active disease display functional impairment of Ag-specific CD8+ T cells due to expansion of regulatory T cells at sites of disease and in the peripheral blood. Adoptive cellular immunotherapy based on EBV-specific CD8+ T cells has been explored with limited success to date. It has been proposed that improved targeting of these CD8+ T cells toward viral Ags that are expressed in HL may enhance future therapeutic vaccine strategies. In this study, we have developed a novel replication-deficient adenoviral Ag presentation system that is designed to encode glycine alanine repeat-deleted EBV nuclear Ag 1 covalently linked to multiple CD8+ T cell epitopes from latent membrane proteins 1 and 2. A single stimulation of CD8+ T cells from healthy virus carriers, and patients with HL with this adenoviral construct in combination with IL-2, was sufficient to reverse the functional T cell impairment and restored both IFN-gamma production and cytolytic function. More importantly, these activated CD8+ T cells responded to tumor cells expressing membrane proteins and recognized novel EBNA1 epitopes. Flow cytometric analysis revealed that a large proportion of T cells expanded from patients with HL were CD62L(high) and CD27(high), and CCR7(low), consistent with early to mid effector T cells. These findings provide an important platform for translation of Ag-specific adoptive immunotherapy for the treatment of EBV-associated malignancies such as HL and nasopharyngeal carcinoma.     

Altered EBV viral load setpoint after HIV seroconversion is in accordance with lack of predictive value of EBV load for the occurrence of AIDS-related non-Hodgkin lymphoma

In contrast to the situation in the post-transplant setting, in HIV-infected individuals an elevated EBV load is not predictive of EBV-related malignancies. To study whether a high EBV load is already a normal situation early in HIV infection and is not related to a decrease in immune function over time, we investigated EBV load and EBV-specific CD8(+) T cells approximately 1 year before and 1 year after HIV seroconversion. EBV load significantly increased after HIV seroconversion from 205 to 1002 copies/10(6) PBMC (p < 0.001), whereas no further increase in EBV load was observed between 1 and 5 years after HIV seroconversion (median, 1827-2478 copies/10(6) PBMC; p = 0.530). Interestingly, the absolute number of EBV lytic epitope, RAKFKQLL-specific CD8(+) T cells increased over HIV seroconversion (4.78 to 9.54/ micro l; p = 0.011). Furthermore, the fraction of CD27-negative effector, RAK-specific CD8(+) T cells tended to increase (from 12.2 to 17.31% CD27(-); p = 0.051), in accordance with Ag-driven differentiation. In conclusion, both virological and immunological data support the idea that a new EBV viral setpoint is reached early in HIV infection, probably by EBV reactivation, as suggested by the preferential increase in EBV lytic epitope-specific CD8(+) T cells. These data may thus help to explain the lack of predictive value of EBV load for the occurrence of AIDS-related lymphoma.     

Preferential apoptosis of HIV-1-specific CD4+ T cells

In contrast to other viral infections such as CMV, circulating frequencies of HIV-1-specific CD4+ T cells in peripheral blood are quantitatively diminished in the majority of HIV-1-infected individuals. One mechanism for this quantitative defect is preferential infection of HIV-1-specific CD4+ T cells, although <10% of HIV-1-specific CD4+ T cells are infected. Apoptosis has been proposed as an important contributor to the pathogenesis of CD4+ T cell depletion in HIV/AIDS. We show here that, within HIV-1-infected individuals, a greater proportion of ex vivo HIV-1-specific CD4+ T cells undergo apoptosis compared with CMV-specific CD4+ T cells (45 vs 7.4%, respectively, p < 0.05, in chronic progressors). The degree of apoptosis within HIV-1-specific CD4+ T cells correlates with viral load and disease progression, and highly active antiretroviral therapy abrogates these differences. The data support a mechanism for apoptosis in these cells similar to that found in activation-induced apoptosis through the TCR, resulting in oxygen-free radical production, mitochondrial damage, and caspase-9 activation. That HIV-1 proteins can also directly enhance activation-induced apoptosis supports a mechanism for a preferential induction of apoptosis of HIV-1-specific CD4+ T cells, which contributes to a loss of immunological control of HIV-1 replication.     

HIV-1-specific memory CD4+ T cells are phenotypically less mature than cytomegalovirus-specific memory CD4+ T cells

HIV-1-specific CD4(+) T cells are qualitatively dysfunctional in the majority of HIV-1-infected individuals and are thus unable to effectively control viral replication. The current study extensively details the maturational phenotype of memory CD4(+) T cells directed against HIV-1 and CMV. We find that HIV-1-specific CD4(+) T cells are skewed to an early central memory phenotype, whereas CMV-specific CD4(+) T cells generally display a late effector memory phenotype. These differences hold true for both IFN-gamma- and IL-2-producing virus-specific CD4(+) T cells, are present during all disease stages, and persist even after highly active antiretroviral therapy (HAART). In addition, after HAART, HIV-1-specific CD4(+) T cells are enriched for CD27(+)CD28(-)-expressing cells, a rare phenotype, reflecting an early intermediate stage of differentiation. We found no correlation between differentiation phenotype of HIV-1-specific CD4(+) T cells and HIV-1 plasma viral load or HIV-1 disease progression. Surprisingly, HIV-1 viral load affected the maturational phenotype of CMV-specific CD4(+) T cells toward an earlier, less-differentiated state. In summary, our data indicate that the maturational state of HIV-1-specific CD4(+) T cells cannot be a sole explanation for loss of containment of HIV-1. However, HIV-1 replication can affect the phenotype of CD4(+) T cells of other specificities, which might adversely affect their ability to control those pathogens. The role for HIV-1-specific CD4(+) T cells expressing CD27(+)CD28(-) after HAART remains to be determined.     

HIV-specific CD8+ T cell function in children with vertically acquired HIV-1 infection is critically influenced by age and the state of the CD4+ T cell compartment

The immunology of vertical HIV transmission differs from that of adult infection in that the immune system of the infant is not fully matured, and the factors that influence the functionality of CD8(+) T cell responses against HIV in children remain largely undefined. We have investigated CD8(+) T cell responses in 65 pediatric subjects with vertically acquired HIV-1 infection. Vigorous, broad, and Ag dose-driven CD8(+) T cell responses against HIV Ags were frequently observed in children who were older than 3 years of age and maintained CD4(+) T cell counts >400 cells/ micro l. In contrast, younger age or a CD4(+) T cell count <400 cells/ micro l was associated with poor CD8(+) T cell responses and high HIV loads. Furthermore, subjects with a severely depleted and phenotypically altered CD4(+) T cell compartment had circulating Gag-specific CD8(+) T cells with impaired IFN-gamma production. When viral load was not suppressed by antiviral treatment, subjects that fell below the putative age and CD4(+) T cell count thresholds had significantly reduced CD8(+) T cell responses and significantly higher viral loads. Thus, the data suggest that fully effective HIV-specific CD8(+) T cell responses take years to develop despite an abundance of Ag in early life, and responses are further severely impaired, independent of age, in children who have a depleted or skewed CD4(+) T cell compartment. The results are discussed in relation to differences between the neonatal and adult immune systems in the ability to respond to HIV infection.     

The CD160+ CD8high cytotoxic T cell subset correlates with response to HAART in HIV-1+ patients

We investigated the circulating cytotoxic CD160+ CD8(high) subset in correlation to antiviral immunity and response to highly active antiretroviral therapy (HAART) in HIV+ subjects. The study included 45 treatment-naive patients receiving HAART for 18 months, retrospectively defined as good (n=29) and transient (n=16) responders. HIV-specific CD8 T lymphocyte levels were measured by IFNgamma production in response to p17 Gag, in the presence of immobilized anti-CD160 mAb. We report a significantly increased baseline level of CD160+ CD8(high) subset in good therapy responders. CD160+ CD8(high) subset correlates with CD4+ T cell count, immune activation, and viral load. CD160+ CD8(high) lymphocytes contain a high amount of Granzyme B and include virus-specific T lymphocytes in HIV-1+ subjects. Co-stimulation through CD160 molecules enhances IFNgamma production in response to p17 Gag. Therefore, the CD160+ CD8(high) subset may be useful for monitoring of virus-specific cellular immunity and predicting response to antiretroviral therapy in chronic HIV-1 infection.     

Viral suppression and immune restoration in the gastrointestinal mucosa of human immunodeficiency virus type 1-infected patients initiating therapy during primary or chronic infection

Although the gut-associated lymphoid tissue (GALT) is an important early site for human immunodeficiency virus (HIV) replication and severe CD4+ T-cell depletion, our understanding is limited about the restoration of the gut mucosal immune system during highly active antiretroviral therapy (HAART). We evaluated the kinetics of viral suppression, CD4+ T-cell restoration, gene expression, and HIV-specific CD8+ T-cell responses in longitudinal gastrointestinal biopsy and peripheral blood samples from patients initiating HAART during primary HIV infection (PHI) or chronic HIV infection (CHI) using flow cytometry, real-time PCR, and DNA microarray analysis. Viral suppression was more effective in GALT of PHI patients than CHI patients during HAART. Mucosal CD4+ T-cell restoration was delayed compared to peripheral blood and independent of the time of HAART initiation. Immunophenotypic analysis showed that repopulating mucosal CD4+ T cells were predominantly of a memory phenotype and expressed CD11 alpha, alpha(E)beta 7, CCR5, and CXCR4. Incomplete suppression of viral replication in GALT during HAART correlated with increased HIV-specific CD8+ T-cell responses. DNA microarray analysis revealed that genes involved in inflammation and cell activation were up regulated in patients who did not replenish mucosal CD4+ T cells efficiently, while expression of genes involved in growth and repair was increased in patients with efficient mucosal CD4+ T-cell restoration. Our findings suggest that the discordance in CD4+ T-cell restoration between GALT and peripheral blood during therapy can be attributed to the incomplete viral suppression and increased immune activation and inflammation that may prevent restoration of CD4+ T cells and the gut microenvironment.     

CD4+ and CD8+ T-Cell Responses to Latent Antigen EBNA-1 and Lytic Antigen BZLF-1 during Persistent Lymphocryptovirus Infection of Rhesus Macaques

Epstein-Barr virus (EBV) infection leads to lifelong viral persistence through its latency in B cells. EBV-specific T cells control reactivations and prevent the development of EBV-associated malignancies in most healthy carriers, but infection can sometimes cause chronic disease and malignant transformation. Epstein-Barr nuclear antigen 1 (EBNA-1) is the only viral protein consistently expressed during all forms of latency and in all EBV-associated malignancies and is a promising target for a therapeutic vaccine. Here, we studied the EBNA-1-specific immune response using the EBV-homologous rhesus lymphocryptovirus (rhLCV) infection in rhesus macaques. We assessed the frequency, phenotype, and cytokine production profiles of rhLCV EBNA-1 (rhEBNA-1)-specific T cells in 15 rhesus macaques and compared them to the lytic antigen of rhLCV BZLF-1 (rhBZLF-1). We were able to detect rhEBNA-1-specific CD4⁺ and/or CD8⁺ T cells in 14 of the 15 animals screened. In comparison, all 15 animals had detectable rhBZLF-1 responses. Most peptide-specific CD4⁺ T cells exhibited a resting phenotype of central memory (TCM), while peptide-specific CD8⁺ T cells showed a more activated phenotype, belonging mainly to the effector cell subset. By comparing our results to the human EBV immune response, we demonstrate that the rhLCV model is a valid system for studying chronic EBV infection and for the preclinical development of therapeutic vaccines.     

Cytolytic CD4(+)-T-cell clones reactive to EBNA1 inhibit Epstein-Barr virus-induced B-cell proliferation

In the absence of immune surveillance, Epstein-Barr virus (EBV)-infected B cells generate neoplasms in vivo and transformed cell lines in vitro. In an in vitro system which modeled the first steps of in vivo immune control over posttransplant lymphoproliferative disease and lymphomas, our investigators previously demonstrated that memory CD4(+) T cells reactive to EBV were necessary and sufficient to prevent proliferation of B cells newly infected by EBV (S. Nikiforow et al., J. Virol. 75:3740-3752, 2001). Here, we show that three CD4(+)-T-cell clones reactive to the latent EBV antigen EBNA1 also prevent the proliferation of newly infected B cells from major histocompatibility complex (MHC) class II-matched donors, a crucial first step in the transformation process. EBNA1-reactive T-cell clones recognized B cells as early as 4 days after EBV infection through an HLA-DR-restricted interaction. They secreted Th1-type and Th2-type cytokines and lysed EBV-transformed established lymphoblastoid cell lines via a Fas/Fas ligand-dependent mechanism. Once specifically activated, they also caused bystander regression and bystander killing of non-MHC-matched EBV-infected B cells. Since EBNA1 is recognized by CD4(+) T cells from nearly all EBV-seropositive individuals and evades detection by CD8(+) T cells, EBNA1-reactive CD4(+) T cells may control de novo expansion of B cells following EBV infection in vivo. Thus, EBNA1-reactive CD4(+)-T-cell clones may find use as adoptive immunotherapy against EBV-related lymphoproliferative disease and many other EBV-associated tumors.     

Dual stimulation of Epstein-Barr Virus (EBV)-specific CD4+- and CD8+-T-cell responses by a chimeric antigen construct: potential therapeutic vaccine for EBV-positive nasopharyngeal carcinoma

Virus-associated malignancies are potential targets for immunotherapeutic vaccines aiming to stimulate T-cell responses against viral antigens expressed in tumor cells. Epstein-Barr virus (EBV)-associated nasopharyngeal carcinoma, a high-incidence tumor in southern China, expresses a limited set of EBV proteins, including the nuclear antigen EBNA1, an abundant source of HLA class II-restricted CD4(+) T-cell epitopes, and the latent membrane protein LMP2, a source of subdominant CD8(+) T-cell epitopes presented by HLA class I alleles common in the Chinese population. We used appropriately modified gene sequences from a Chinese EBV strain to generate a modified vaccinia virus Ankara recombinant, MVA-EL, expressing the CD4 epitope-rich C-terminal domain of EBNA1 fused to full-length LMP2. The endogenously expressed fusion protein EL is efficiently processed via the HLA class I pathway, and MVA-EL-infected dendritic cells selectively reactivate LMP2-specific CD8(+) memory T-cell responses from immune donors in vitro. Surprisingly, endogenously expressed EL also directly accesses the HLA class II presentation pathway and, unlike endogenously expressed EBNA1 itself, efficiently reactivates CD4(+) memory T-cell responses in vitro. This unscheduled access to the HLA class II pathway is coincident with EL-mediated redirection of the EBNA1 domain from its native nuclear location to dense cytoplasmic patches. Given its immunogenicity to both CD4(+) and CD8(+) T cells, MVA-EL has potential as a therapeutic vaccine in the context of nasopharyngeal carcinoma.     

Human Cytomegalovirus Latency-Associated Proteins Elicit Immune-Suppressive IL-10 Producing CD4+ T Cells

Human cytomegalovirus (HCMV) is a widely prevalent human herpesvirus, which, after primary infection, persists in the host for life. In healthy individuals, the virus is well controlled by the HCMV-specific T cell response. A key feature of this persistence, in the face of a normally robust host immune response, is the establishment of viral latency. In contrast to lytic infection, which is characterised by extensive viral gene expression and virus production, long-term latency in cells of the myeloid lineage is characterised by highly restricted expression of viral genes, including UL138 and LUNA. Here we report that both UL138 and LUNA-specific T cells were detectable directly ex vivo in healthy HCMV seropositive subjects and that this response is principally CD4⁺ T cell mediated. These UL138-specific CD4⁺ T cells are able to mediate MHC class II restricted cytotoxicity and, importantly, show IFNγ effector function in the context of both lytic and latent infection. Furthermore, in contrast to CD4⁺ T cells specific to antigens expressed solely during lytic infection, both the UL138 and LUNA-specific CD4⁺ T cell responses included CD4⁺ T cells that secreted the immunosuppressive cytokine cIL-10. We also show that cIL-10 expressing CD4⁺ T-cells are directed against latently expressed US28 and UL111A. Taken together, our data show that latency-associated gene products of HCMV generate CD4⁺ T cell responses in vivo, which are able to elicit effector function in response to both lytic and latently infected cells. Importantly and in contrast to CD4⁺ T cell populations, which recognise antigens solely expressed during lytic infection, include a subset of cells that secrete the immunosuppressive cytokine cIL-10. This suggests that HCMV skews the T cell responses to latency-associated antigens to one that is overall suppressive in order to sustain latent carriage in vivo.     

IL-15-independent proliferative renewal of memory CD8+ T cells in latent gammaherpesvirus infection

IL-15 is known to be critical in the homeostasis of Ag-specific memory CD8(+) T cells following acute viral infection. However, little is known about the homeostatic requirements of memory CD8(+) T cells during a latent viral infection. We have used the murine gammaherpesvirus-68 (MHV-68) model system to investigate whether IL-15 is necessary for the maintenance of memory CD8(+) T cells during a latent viral infection. IL-15 is not essential either for the initial control of MHV-68 infection or for the maintenance of MHV-68-specific memory CD8(+) T cells. Even at 140 days postinfection, the proportion of CD8(+) T cells recognizing the MHV-68 epitopes were the same as in control mice. The maintenance of these memory CD8(+) T cells was attributable to their ability to turn over in vivo, probably in response to the presence of low levels of Ag. IL-15(-/-) mice had a significantly higher turnover rate within the virus-specific memory CD8(+) T cell population, which was the result of increased levels of viral gene expression rather than an increase in viral load. These cells did not accumulate in the spleens of the IL-15(-/-) mice due to an increased sensitivity to apoptosis as a result of decreased Bcl-2 levels. Intriguingly, memory CD8(+) T cells from latently infected mice failed to undergo homeostatic proliferation in a naive secondary host. These data highlight fundamental differences between memory CD8(+) T cells engaged in active immune surveillance of latent viral infections vs memory CD8(+) T cells found after acute viral infections.     

Residual Viral Replication during Antiretroviral Therapy Boosts Human Immunodeficiency Virus Type 1-Specific CD8+ T-Cell Responses in Subjects Treated Early after Infection

Human immunodeficiency virus type 1 (HIV-1)-infected subjects treated early after infection have preserved HIV-1-specific CD4+ T-cell function. We studied the effect of highly active antiretroviral therapy (HAART) on the frequency of HIV-1-specific CD8+ T cells in patients treated during early (n = 31) or chronic (n = 23) infection. The degree of viral suppression and time of initiation of treatment influenced the magnitude of the CD8+ T-cell response. HIV-1-specific CD8+ T cells can increase in number after HAART in subjects treated early after infection who have episodes of transient viremia.     

CD40 ligand trimer and IL-12 enhance peripheral blood mononuclear cells and CD4+ T cell proliferation and production of IFN-gamma in response to p24 antigen in HIV-infected individuals: potential contribution of anergy to HIV-specific unresponsiveness

It has been suggested that CD4+ T cell proliferative responses to HIV p24 Ag may be important in the control of HIV infection. However, these responses are minimal or absent in many HIV-infected individuals. Furthermore, while in vitro and in vivo responses to non-HIV recall Ags improve upon administration of highly active antiretroviral therapy, there does not appear to be a commensurate enhancement of HIV-specific immune responses. It is possible that CD4+ p24-specific T cells are deleted early in the course of infection. However, it is also possible that a discrete unresponsiveness, or anergy, contributes to the lack of proliferation to p24. To evaluate the possible contribution of unresponsiveness to the lack of CD4+ T cell proliferation to p24 in HIV-infected individuals, we attempted to overcome unresponsiveness. CD40 ligand trimer (CD40LT) and IL-12 significantly increased PBMC and CD4+ T cell proliferative responses to p24 Ag in HIV-infected, but not uninfected, individuals. No increase in proliferative response to CMV Ag was observed. CD40LT exerted its effect through B7-CD28-dependent and IL-12- and IL-15-independent mechanisms. Finally, the increase in proliferation with CD40LT and IL-12 was associated with an augmented production of IFN-gamma in most, but not all, individuals. These data suggest the possible contribution of HIV-specific unresponsiveness to the lack of CD4+ T cell proliferation to p24 Ag in HIV-infected individuals and that clonal deletion alone does not explain this phenomenon. They also indicate the potential for CD40LT and IL-12 as immune-based therapies for HIV infection.