Dissecting the host response to a gamma-herpesvirus

The murine gamma-herpesvirus 68 (MHV-68) provides a unique experimental model for dissecting immunity to large DNA viruses that persist in B lymphocytes. The analysis is greatly facilitated by the availability of genetically disrupted (-/-) mice that lack key host-response elements, and by the fact that MHV-68 is a lytic virus that can readily be manipulated for mutational analysis. The mutant virus strategy is being used, for example, to characterize the part played in vivo by an MHV-68-encoded chemokine-binding protein that may ultimately find an application in human therapeutics. Experiments with various -/- mice and monoclonal antibody depletion protocols have shown very clearly that type I interferons (IFNs) are essential for the early control of MHV-68 replication, while CD4+ T cells producing IFN-gamma function to limit the consequences of viral persistence. Virus-specific CD8+ effectors acting in the absence of the CD4+ subset seem initially to control the lytic phase in the lung following respiratory challenge, but are then unable to prevent the reactivation of replicative infection in epithelia and the eventual death of CD4+ T-cell-deficient mice. This could reflect the fact that the interaction between the CD8+ T cells and the virus-infected targets is partially compromised by the MHV-68 K3 protein, which inhibits antigen presentation by MHC class I glycoproteins. Immunization strategies focusing on the CD8+ T-cell response to epitopes expressed during the lytic phase of MHV-68 infection can limit virus replication, but are unable to prevent the establishment of latency. Other experiments with mutant viruses also suggest that there is a disconnection between lytic MHV-68 infection and latency. The massive nonspecific immunoglobulin response and the dramatic expansion of Vbeta4+ CD8+ T cells, which is apparently MHC independent, could represent some sort of 'smoke screen' used by MHV-68 to subvert immunity. Although MHV-68 is neither Epstein-Barr virus nor human herpesvirus-8, the results generated from this system suggest possibilities that may usefully be addressed with these human pathogens. Perhaps the main lesson learned to date is that all the components of immunity are likely to be important for the control of these complex viruses.     

CD8(+) T cells from mice transnuclear for a TCR that recognizes a single H-2K(b)-restricted MHV68 epitope derived from gB-ORF8 help control infection

To study the CD8(+) T cell response against a mouse γ-herpes virus, we generated K(b)-MHV-68-ORF8(604-612)RAG(-/-) CD8(+) T cell receptor transnuclear (TN) mice as a source of virus-specific CD8(+) T cells. K(b)-ORF8-Tet(+) CD8(+) T cells, expanded in the course of a resolving MHV-68 infection, served as a source of nucleus donors. Various in vivo and ex vivo assay criteria demonstrated the fine specificity and functionality of TN cells. TN cells proliferated extensively in response to viral infection, helped control viral burden, and exhibited a phenotype similar to that of endogenous K(b)-ORF8-Tet(+) cells. When compared to OT-1 cells, TN cells displayed distinct properties in response to lymphopenia and cognate antigen stimulation, which may be attributable to the affinity of the TCR expressed by the TN cells. The availability of MHV-68-specific CD8(+) TCR TN mice provides a new tool for investigating aspects of host-pathogen interactions unique to γ-herpes viruses.     

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.     

Apparent MHC-independent stimulation of CD8+ T cells in vivo during latent murine gammaherpesvirus infection.

Like EBV-infected humans with infectious mononucleosis, mice infected with the rodent gammaherpesvirus MHV-68 develop a profound increase in the number of CD8+ T cells in the circulation. In the mouse model, this lymphocytosis consists of highly activated CD8+ T cells strikingly biased toward V beta 4 TCR expression. Moreover, this expansion of V beta 4+CD8+ T cells does not depend on the MHC haplotype of the infected animal. Using a panel of lacZ-inducible T cell hybridomas, we have detected V beta 4-specific T cell stimulatory activity in the spleens of MHV-68-infected mice. We show that the appearance and quantity of this activity correlate with the establishment and magnitude of latent viral infection. Furthermore, on the basis of Ab blocking studies as well as experiments with MHC class II, beta2-microglobulin (beta2m) and TAP1 knockout mice, the V beta 4-specific T cell stimulatory activity does not appear to depend on conventional presentation by classical MHC class I or class II molecules. Taken together, the data indicate that during latent infection, MHV-68 may express a T cell ligand that differs fundamentally from both conventional peptide Ags and classical viral superantigens.     

Protein kinase C theta is not essential for T-cell-mediated clearance of murine gammaherpesvirus 68

Murine gammaherpesvirus 68 (MHV-68) is a naturally occurring rodent pathogen with significant homology to human pathogens Epstein-Barr virus and Kaposi's sarcoma-associated herpesvirus. T cells are essential for primary clearance of MHV-68 and survival of mice following intranasal infection. Previous reports have suggested that protein kinase C theta (PKCtheta) is essential for T-cell activation and cytokine production in vitro. To determine the role of this molecule in vivo during the immune response to a viral infection, PKCtheta-/- mice were infected with MHV-68. Despite the essential role of T cells in viral clearance, PKCtheta-/- mice survived infection, cleared lytic virus, and maintained effective long-term control of latency. CD8 T-cell expansion, trafficking to the lung, and cytotoxic activity were similar in PKCtheta+/+ and PKCtheta-/- mice, whereas antiviral antibody and T-helper cell cytokine production were significantly lower in PKCtheta-/- mice than in PKCtheta+/+ mice. These studies demonstrate a differential requirement for PKCtheta in the immune response to MHV-68 and show that PKCtheta is not essential for the T-cell activation events leading to viral clearance.     

CD8+ T cell dysfunction and increase in murine gammaherpesvirus latent viral burden in the absence of 4-1BB ligand

Studies of costimulatory receptors belonging to the TNFR family have revealed their diverse roles in affecting different stages of the T cell response. The 4-1BB ligand (4-1BBL)/4-1BB pathway has emerged as a receptor-ligand pair that impacts not the initial priming, but later phases of the T cell response, such as sustaining clonal expansion and survival, maintaining memory CD8(+) T cells, and supporting secondary expansion upon Ag challenge. Although the role of this costimulatory pathway in CD8(+) T cell responses to acute viral infections has been well-studied, its role in controlling chronic viral infections in vivo is not known to date. Using the murine gammaherpesvirus-68 (MHV-68) model, we show that 4-1BBL-deficient mice lack control of MHV-68 during latency and show significantly increased latent viral loads. In contrast to acute influenza infection, the numbers of MHV-68-specific memory CD8(+) T cells were maintained during latency. However, the virus-specific CD8(+) T cells showed defects in function, including decreased cytolytic function and impaired secondary expansion. Thus, 4-1BBL deficiency significantly affects the function, but not the number, of virus-specific CD8(+) T cells during gammaherpesvirus latency, and its absence results in an increased viral burden. Our study suggests that the 4-1BB costimulatory pathway plays an important role in controlling chronic viral infections.     

Protective vaccination with hepatitis C virus NS3 but not core antigen in a novel mouse challenge model

BACKGROUND: Efficient vaccines against hepatitis C virus (HCV) infection are urgently needed. Vaccine development has been hampered by the lack of suitable small animal models to reliably test the protective capacity of immmunization. METHODS: We used recombinant murine gammaherpesvirus 68 (MHV-68) as a novel challenge virus in mice and tested the efficacy of heterologous candidate human vaccines based on modified vaccinia virus Ankara or adenovirus, both delivering HCV non-structural NS3 or core proteins. RESULTS: Recombinant MHV-68 expressing NS3 (MHV-68-NS3) or core (MHV-68-core) were constructed and characterized in vitro and in vivo. Mice immunized with NS3-specific vector vaccines and challenged with MHV-68-NS3 were infected but showed significantly reduced viral loads in the acute and latent phase of infection. NS3-specific CD8+ T cells were amplified in immunized mice after challenge with MHV-68-NS3. By contrast, we did neither detect a reduction of viral load nor an induction of core-specific CD8+ T cells after core-specific immunization. CONCLUSIONS: Our data suggest that the challenge system using recombinant MHV-68 is a highly suitable model to test the immunogenicity and protective capacity of HCV candidate vaccine antigens. Using this system, we demonstrated the usefulness of NS3-specific immunization. By contrast, our analysis rather discarded core as a vaccine antigen.     

The CD8 T-cell response against murine gammaherpesvirus 68 is directed toward a broad repertoire of epitopes from both early and late antigens

Infection of mice with murine gammaherpesvirus 68 (MHV-68) robustly activates CD8 T cells, but only six class I major histocompatibility complex (MHC)-restricted epitopes have been described to date for the widely used H-2(b) haplotype mice. To explore the specificity and kinetics of the cytotoxic T-lymphocyte response in MHV-68-infected C57BL/6 mice, we screened for H-2K(b)- and H-2D(b)-restricted epitopes using a set of 384 candidate epitopes in an MHC tetramer-based approach and identified 19 new epitopes in 16 different open reading frames. Of the six known H-2K(b)- and H-2D(b)-restricted epitopes, we confirmed a response against three and did not detect CD8 T-cell-specific responses for the remaining three. The peak of the CD8 T-cell response to most peptides occurs between 6 and 10 days postinfection. The respective MHC tetramer-positive CD8 T cells display an activated/effector phenotype (CD62L(lo) and CD44(hi)) and produce gamma interferon upon peptide stimulation ex vivo. MHV-68 infection in vivo elicits a response to multiple viral epitopes, derived from both early and late viral antigens, illustrating a far broader T-cell repertoire and more-rapid activation than those previously recorded.     

Visualization of polyoma virus-specific CD8+ T cells in vivo during infection and tumor rejection.

T cells are critical for clearing infection and preventing tumors induced by polyoma virus, a natural murine papovavirus. We previously identified the immunodominant epitope for polyoma virus-specific CTL in tumor-resistant H-2k mice as the Dk-restricted peptide, MT389-397, derived from the polyoma middle T oncoprotein. In this study, we developed tetrameric Dk complexes containing the MT389-397 peptide to directly visualize and enumerate MT389-397-specific CTL during polyoma virus infection. We found that Dk/MT389 tetramer+CD8+ T cells undergo a massive expansion during primary infection such that by day 7 postinfection these Ag-specific CD8+ T cells constitute approximately 20% of the total and approximately 40% of the activated CD8+ T cells in the spleen. This expansion of Dk/MT389 tetramer+CD8+ T cells parallels the emergence of MT389-397-specific ex vivo cytolytic activity and clearance of polyoma virus. Notably, Dk/MT389 tetramer+CD8+ T cells are maintained in memory at very high levels. The frequencies of Dk/MT389 tetramer+CD8+ effector and memory T cells in vivo match those of CD8+ T cells producing intracellular IFN-gamma after 6-h in vitro stimulation by MT389-397 peptide. Consistent with preferential Vbeta6 expression by MT389-397-specific CD8+CTL lines and clones, Dk/MT389 tetramer+CD8+ T cells exhibit biased expression of this Vbeta gene segment. Finally, we show that Dk/MT389 tetramer+CD8+ T cells efficiently infiltrate a polyoma tumor challenge to virus-immune mice. Taken together, these findings strongly implicate virus-induced MT389-397-specific CD8+ T cells as essential effectors in eliminating polyoma-infected and polyoma-transformed cells in vivo.     

Antiviral protection by CD8+ versus CD4+ T cells. CD8+ T cells correlating with cytotoxic activity in vitro are more efficient in antivaccinia virus protection than CD4-dependent IL.

T cell-mediated protection against a recombinant vaccinia virus was evaluated in mice with respect to the relative contributions of CTL vs that of T cell-dependent IL and of CD4+ cells. H-2b mice primed with the wildtype of vesicular stomatitis virus serotype Indiana (VSV-IND wt) mount an in vitro measurable cytotoxic response against the nucleoprotein (NP) of VSV-IND and are protected against a challenge infection with a vaccinia-VSV recombinant virus expressing the NP of VSV-IND (vacc-IND-NP). Their protective mechanism was highly susceptible to in vivo depletion of CD8+ T cells, but resistant to CD4+ depletion or treatment with anti-IFN-gamma and anti-TNF-alpha. Surprisingly, also VSV-CTL nonresponder H-2k mice were protected against a challenging infection with vacc-IND-NP when primed with VSV-IND wt. In contrast to the CTL responder H-2b mice, this protection was highly susceptible to CD4+ T cell depletion and to anti-IFN-gamma or anti-TNF-alpha treatment, but resistant to CD8+ T cell depletion. Antibodies were not responsible because they failed to transfer protection; in contrast CD4+ T cells conferred significant protection. VSV-CTL responder H-2b and nonresponder H-2k mice were protected almost equally well against a challenge dose of 10(3) pfu vacc-IND-NP inoculated intracerebrally. However, after intracerebral challenge with 5 x 10(6) pfu vacc-IND-NP, the CTL nonresponder mice died, whereas the CTL responder mice eliminated the virus by day 5. These results collectively show that CD4+ T cell-dependent IL may mediate antiviral protection, but their efficiency is relatively weak compared with CD8-mediated protection correlating with cytotoxic activity in vitro.     

A Primary Study of the Subgroups of T Lymphocytes in MHV-3 Induced Chronic Viral Hepatitis

To study the contribution of T cell subsets in the pathogenesis of Murine hepatitis virus Type 3(MHV-3) induced chronic viral hepatitis in C3H/Hej mice,ninety C3H/Hej mice were chosen to individually receive 10 plaque forming units(PFU)of MHV-3 intraperitoneally.The changes of virus titer and pathology in liver tissue were examined by standard plaque assay and by the hematoxylin/eosin(HE) staining method from 2 days post MHV-3 infection.The ratios of T cell subsets including CD3 CD4 CD8-,CD3 CD4-CD8 ,CD3 CD4-CD8-,CD3 CD4 CD25 ,CD3 CD4 CD25-and CD3 CD4-CD25 T lymphocyte of total T lymphocytes in blood,spleen and liver were examined at 0,2,4,6,8,10,12,15,20,25,30,40 days post MHV-3 infection by flow cytosorting.We observed that the virus titer raised and showed persistent virus duplications and inflammatory changes in the livers of C3H/Hej mice from 2 days post MHV-3 infection.The double negative T cell(DN Treg cell) and CD4 CD25 T cell ratios increased significantly from 2 days post MHV-3 infection in C3H/Hej mice,and CD3 CD4 CD8-,CD3 CD4-CD8 ,CD3 CD4 CD25-and CD3 CD4-CD25 T cell ratios decreased accordingly.In conclusion,the changes of virus titer and pathology in the livers of C3H/Hej mice post MHV-3 suggest their contribution to viral persistence.Further characterizations of DN Treg cells are that infection indicates that MHV-3 could induce the chronic inflammation in livers of C3H/Hej mice.The increase of the DN Treg cell and CD4 CD25 T cell ratios in C3H/Hej mice post MHV-3 infection suggests that DN Treg cells and CD4 CD25 T cells may both have important suppressive immunomodulation functions in the development of chronic viral hepatitis and have important roles in the virus persistent infection.Further characterizations of DNT cell and CD4 CD25 T cell are under investigation.     

CD8+ T cells are a biologically relevant source of macrophage inflammatory protein-1 alpha in vivo

Chemokines are small proteins that direct the migration of leukocytes to inflammatory foci. Many cell types, including macrophages, fibroblasts, endothelial cells, and lymphocytes, produce chemokines in vitro, but biologically relevant sources of chemokines in vivo have not been well characterized. To investigate the pertinent sources of macrophage inflammatory protein-1 alpha (MIP-1 alpha) in vivo, we used MIP-1 alpha-deficient (MIP-1 alpha-/-) mice as donors and as recipients in adoptive transfer experiments after a lethal infection with Listeria monocytogenes (LM). Unexpectedly, we found that the production of MIP-1 alpha by CD8+ T cells was critical in this system, as the cells from MIP-1 alpha-/- mice primed with LM were significantly less effective in protecting naive mice against a lethal infection by LM than were the CD8+ T cells from wild-type (wt) mice. This requirement for donor T cell production of MIP-1 alpha was confirmed by the observation that wt donor T cells do not mediate protection when coadministered with an anti-MIP-1 alpha polyclonal antiserum. Production of MIP-1 alpha by the recipient mice was not required for protection, because wt and MIP-1 alpha-/- recipients were equally well protected by wt T cells. A 2- to 3-fold decrease in the number of transferred lymphocytes was seen in the spleens of mice receiving T cells from MIP-1 alpha-/- mice compared with those receiving wt T cells. In addition, CD8+ T cells from MIP-1 alpha-/- mice had a reduced ability to kill LM-infected target cells in vitro. These findings demonstrate that T cell production of MIP-1 alpha is required for clearance of an intracellular pathogen in vivo.     

Control of memory CD8+ T cell differentiation by CD80/CD86-CD28 costimulation and restoration by IL-2 during the recall response

Memory CD8+ T cell responses have been considered to be independent of CD80/CD86-CD28 costimulation. However, recall responses are often severely blunted in CD28-/- mice. Whether this impairment represents a requirement for CD28 costimulation for proper memory CD8+ T cell development or a requirement during the recall response is unknown. Furthermore, how CD28 costimulation affects the phenotype and function of memory CD8+ T cells has not been characterized in detail. In this study, we investigate these questions by studying the role of the CD28 costimulatory pathway in memory CD8+ T cell responses to acute and persistent DNA virus infections. Memory CD8+ T cells against vaccinia virus (VV) infection which develop without CD28 costimulation exhibit lower expression of differentiation markers CD27 and CD122 (IL-15Rbeta). These memory CD8+ T cells also fail to produce IL-2. Our data indicate that for an optimal recall response, CD28 costimulation is required both for T cell priming and also during the recall response. Similar requirements were observed for memory CD8+ T cell responses during persistent infection with murine gammaherpesvirus 68 (MHV-68) infection, indicating CD28 may play the same role in both acute and persistent infections. Finally, we show deficits in the recall response are restored by IL-2 signaling during recall, but not during priming. The data presented show that CD28 costimulation not only controls the magnitude of the primary response but also affects development of memory CD8+ T cells and is required during the recall response in addition to initial T cell priming.     

Latent murine gamma-herpesvirus infection is established in activated B cells, dendritic cells, and macrophages

Intranasal infection of mice with the murine gamma-herpesvirus MHV-68 results in an acute lytic infection in the lung, followed by the establishment of lifelong latency. Development of an infectious mononucleosis-like syndrome correlates with the establishment of latency and is characterized by splenomegaly and the appearance of activated CD8+ T cells in the peripheral blood. Interestingly, a large population of activated CD8+ T cells in the peripheral blood expresses the V beta 4+ element in their TCR. In this report we show that MHV-68 latency in the spleen after intranasal infection is harbored in three APC types: B cells, macrophages, and dendritic cells. Surprisingly, since latency has not previously been described in dendritic cells, these cells harbored the highest frequency of latent virus. Among B cells, latency was preferentially associated with activated B cells expressing the phenotype of germinal center B cells, thus formally linking the previously reported association of latency gene expression and germinal centers to germinal center B cells. Germinal center formation, however, was not required for the establishment of latency. Significantly, although three cell types were latently infected, the ability to stimulate V beta 4+CD8+ T cell hybridomas was limited to latently infected, activated B cells.     

CD40 engagement on dendritic cells, but not on B or T cells, is required for long-term control of murine gammaherpesvirus 68

CD4 T cells are not essential for primary clearance of replicating murine gammaherpesvirus 68 (MHV-68) but are required for effective long-term control. The virus reactivates in the lungs of major histocompatibility complex class II-deficient (CII-/-) mice that lack functional CD4 T cells. CD40 ligand (CD40L) is upregulated on activated CD4 T cells, and it is thought that CD40-CD40L interactions are an important component of CD4 T-cell help. Our previous studies have shown that agonistic antibodies to CD40 can substitute for CD4 T-cell function in the long-term control of MHV-68. In the present study, we sought to identify the CD40-positive cell type mediating this effect. To address this question, we adoptively transferred MHV-68 peptide-pulsed CII(-/-) dendritic cells (DC) that had been treated with an agonistic antibody to CD40 into MHV-68-infected CII(-/-) recipients. Viral reactivation was significantly lower in mice injected with anti-CD40-treated DC than in those injected with control DC or in mice that did not receive any DC. However, in similar experiments with B cells, anti-CD40 treatment had no effect. We also investigated the requirement for CD40 expression on T cells by adoptive transfer of T cells from CD40(+/+) or CD40(-/-) mice into T-cell-deficient recipients that were subsequently infected with MHV-68. The results showed that CD40 expression on T cells is not necessary for preventing viral reactivation. Taken together, our data suggest that CD40 engagement on DC, but not on T or B cells, is essential for effective long-term control of MHV-68.     

Antigen-specific CD8+ T cells and the development of central memory during Mycobacterium tuberculosis infection

Whether true memory T cells develop in the face of chronic infection such as tuberculosis remains controversial. To address this question, we studied CD8+ T cells specific for the Mycobacterium tuberculosis ESAT6-related Ags TB10.3 and TB10.4. The shared epitope TB10.3/10.4(20-28) is presented by H-2 K(d), and 20-30% of the CD8+ T cells in the lungs of chronically infected mice are specific for this Ag following respiratory infection with M. tuberculosis. These TB10.3/10.4(20-28)-specific CD8+ T cells produce IFN-gamma and TNF and express CD107 on their cell surface, which indicates their likely role as CTL in vivo. Nearly all of the Ag-specific CD8+ T cells in the lungs of chronically infected mice had a T effector cell phenotype based on their low expression of CD62L and CD45RB. In contrast, a population of TB10.3/10.4(20-28)-specific CD8+ T cells was identified in the lymphoid organs that express high levels of CD62L and CD45RB. Antibiotic treatment to resolve the infection led to a contraction of the Ag-specific CD8+ T cell population and was accompanied by an increase in the proportion of CD8+ T cells with a central memory phenotype. Finally, challenge of memory-immune mice with M. tuberculosis was accompanied by significant expansion of TB10.3/10.4(20-28)-specific CD8+ T cells, which suggests that these cells are in fact functional memory T cells.     

Ly9 (CD229)-deficient mice exhibit T cell defects yet do not share several phenotypic characteristics associated with SLAM- and SAP-deficient mice

Signaling lymphocyte activation molecule (SLAM) family receptors are critically involved in modulating innate and adaptive immune responses. Several SLAM family receptors have been shown to interact with the adaptor molecule SAP; however, subsequent intracellular signaling is poorly defined. Notably, mutations in SLAM-associated protein (SAP) lead to X-linked lymphoproliferative disease, a rare but fatal immunodeficiency. Although the SLAM family member Ly9 (CD229) is known to interact with SAP, the functions of this receptor have remained elusive. Therefore, we have generated Ly9-/- mice and compared their phenotype with that of SLAM-/- and SAP-/- mice. We report that Ly9-/- T cells exhibit a mild Th2 defect associated with reduced IL-4 production after stimulation with anti-TCR and anti-CD28 in vitro. This defect is similar in magnitude to the previously reported Th2 defect in SLAM-/- mice but is more subtle than that observed in SAP-/- mice. In contrast to SLAM-/- and SAP-/- mice, T cells from Ly9-/- mice proliferate poorly and produce little IL-2 after suboptimal stimulation with anti-CD3 in vitro. We have also found that Ly9-/- macrophages exhibit no defects in cytokine production or bacterial killing as was observed in SLAM-/- macrophages. Additionally, Ly9-/- mice differ from SAP-/- mice in that they foster normal development of NKT cells and mount appropriate T and B cell responses to lymphocytic choriomeningitis virus. We have identified significant phenotypic differences between Ly-9-/- mice as compared with both SLAM-/- and SAP-/- mice. Although Ly9, SLAM, and SAP play a common role in promoting Th2 polarization, Ly-9 is uniquely involved in enhancing T cell activation.     

Suppressive CD8+ T cells arise in the absence of CD4 help and compromise control of persistent virus

There is an urgent need to develop novel therapies for controlling chronic virus infections in immunocompromised patients. Disease associated with persistent γ-herpesvirus infection (EBV, human herpesvirus 8) is a significant problem in AIDS patients and transplant recipients, and clinical management of these conditions is difficult. Immune surveillance failure followed by γ-herpesvirus recrudescence can be modeled using murine γ-herpesvirus (MHV)-68 in mice lacking CD4(+) T cells. In contrast with other chronic infections, no obvious defect in the functional capacity of the viral-specific CD8(+) T cell response was detected. We show in this article that adoptive transfer of MHV-68-specific CD8(+) T cells was ineffective at reducing the viral burden. Together, these indicate the potential presence of T cell extrinsic suppressive factors. Indeed, CD4-depleted mice infected with MHV-68 express increased levels of IL-10, a cytokine capable of suppressing the function of both APCs and T cells. CD4-depleted mice developed a population of CD8(+) T cells capable of producing IL-10 that suppressed viral control. Although exhibiting cell surface markers indicative of activation, the IL-10-producing cells expressed increased levels of programmed death-1 but were not enriched in the MHV-68-specific compartment, nor were they uniformly CD44(hi). Therapeutic administration of an IL-10R blocking Ab enhanced control of the recrudescent virus. These data implicate IL-10 as a promising target for the restoration of immune surveillance against chronic γ-herpesvirus infection in immunosuppressed individuals.     

Augmentation of the CD8+ T cell response by IFN-gamma in IL-12-deficient mice during Toxoplasma gondii infection

The importance of IFN-gamma in regulating the host CD8+ T cell response during microbial infection has not been delineated. Mice deficient for the p40 chain of the IL-12 heterodimer have impaired IFN-gamma production and are susceptible to infection with the intracellular parasite Toxoplasma gondii. The administration of exogenous IFN-gamma to parasite-infected p40-/- mice increases survival and up-regulates the depressed CD8+ T cell response following infection. CD8+ T cells isolated from cytokine-treated p40-/- mice exhibit an increase in both precursor CTL frequency and IFN-gamma production compared with untreated controls. The enhancement of the CD8+ T cell response is independent of CD4+ T cell help. These CD8+ T cells induce protective immunity against a lethal challenge when adoptively transferred into naive p40-/- and IFN-gamma-/- mice. These observations indicate that IFN-gamma can regulate the CD8+ T cell response during T. gondii infection.     

Delayed clearance of Sendai virus in mice lacking class I MHC-restricted CD8+ T cells.

The role and interdependence of CD8+ and CD4+ alpha beta-T cells in the acute response after respiratory infection with the murine parainfluenza type 1 virus, Sendai virus, has been analyzed for H-2b mice. Enrichment of CD8+ virus-specific CTL effectors in the lungs of immunologically intact C57BL/6 animals coincided with the clearance of the virus from this site by day 10 after infection. Removal of the CD4+ T cells by in vivo mAb treatment did not affect appreciably either the recruitment of CD8+ T cells to the infected lung, or their development into virus-specific cytotoxic effectors. In contrast, depletion of the CD8+ subset delayed virus clearance, although most mice survived the infection. Transgenic H-2b F3 mice homozygous (-/-) for a beta 2 microglobulin (beta 2-m) gene disruption, which lack both class I MHC glycoproteins and mature CD8+ alpha beta-T cells, showed a comparable, delayed clearance of Sendai virus from the lung. Virus-specific, class II MHC-restricted CTL were demonstrated in both freshly isolated bronchoalveolar lavage populations and cultured lymph node and spleen tissue from the beta 2-m (-/-) transgenics. Treatment of the beta 2-m (-/-) mice with the mAb to CD4 led to delayed virus clearance and death, which was also the case for normal mice that were depleted simultaneously of the CD4+ and CD8+ subsets. These results indicate that, although classical class I MHC-restricted CD8+ cytotoxic T cells normally play a dominant role in the recovery of mice acutely infected with Sendai virus, alternative mechanisms involving CD4+ T cells exist and can compensate, in time, for the loss of CD8+ T cell function.