Identification of Trypanosoma cruzi trans-sialidase family members as targets of protective CD8+ TC1 responses

CD8+ T lymphocytes play a critical role in immunity to Trypanosoma cruzi. However, the target molecules of this T cell subset have not been elucidated. In this work, we report the identification of an H-2Kb-restricted CTL epitope within two trypomastigote surface Ags encoded by members of the T. cruzi sialidase/trans-sialidase gene superfamily. Octapeptide VDYNFTIV sensitized target cells for lysis by CD8+ CTL generated from spleens of T. cruzi-infected mice. Peptide-specific CD8+ T cell lines were cytotoxic, secreted IFN-gamma and TNF-alpha, but low to undetectable levels of IL-4 and IL-5, and were able, upon adoptive transfer, to confer a high degree of protection against challenge infection. Finally, the protective determinant appears to be conserved among parasites from diverse geographic locations. This constitutes the first identified class I MHC-restricted epitope in T. cruzi and provides the basis for the search of additional targets to be considered in the development of vaccines against Chagas' disease.     

Influence of acute-phase parasite load on pathology, parasitism, and activation of the immune system at the late chronic phase of Chagas' disease

To obtain low and high parasite loads in the acute phase of Chagas' disease, A/J mice were infected with 10(3) or 10(5) Trypanosoma cruzi trypomastigotes of the Y strain and treated on day 6 with benznidazol. One year later, chronically infected mice were screened for subpatent parasitemias, tissue pathology, and immune response. Mice infected with the high parasite inoculum showed higher levels of chronic parasitemias, heart and striated muscle inflammation, and activation of the immune system than did mice infected with the low inoculum. Concerning the activation of the immune system, the main findings for high-dose-infected mice were (i) increased numbers of splenocytes, with preferential expansion of CD8(+) and B220(-) CD5(-) cells, many of them bearing a macrophage phenotype; (ii) higher frequencies of B (B220(+)), CD4(+), and CD8(+) large lymphocytes; (iii) a shift of CD4(+) cells towards a CD45RBLow phenotype; (iv) increased frequencies of both CD45RBLow and CD45RBHigh large CD4(+) cells; (v) augmented numbers of total immunoglobulin (Ig)-secreting cells, with predominance of IgG2a-producing cells; and (vi) increased production of gamma interferon and interleukin 4. In addition, these mice presented lower IgM and higher IgG2a and IgG1 parasite-specific serum antibody levels. Our results indicate that the parasite load at the acute phase of T. cruzi infection influences the activation of the immune system and development of Chagas' disease pathology at the late chronic phase of the disease.     

Physical and functional association between thymic shared antigen-1/stem cell antigen-2 and the T cell receptor complex

Thymic shared antigen-1 (TSA-1)/stem cell Ag-2 (Sca-2) is a glycosylphosphatidylinositol (GPI)-anchored antigen expressed on lymphocytes. We have previously demonstrated that a signal via TSA-1/Sca-2 inhibits T cell receptor (TCR)-mediated T cell activation and apoptosis. To elucidate a molecular mechanism for TSA-1-mediated modulation of the TCR-signaling pathway, we examined whether TSA-1 is physically coupled to the TCR in the present study. TSA-1 was clearly associated with CD3zeta chains in T cell hybridomas, activated T cells, and COS-7 cells transfected with TSA-1 and CD3zeta cDNA. The physical association was confirmed on the surface of T cells in immunoprecipitation and confocal microscopy. The analysis using stable and transient transfectants expressing a transmembrane form of TSA-1 revealed that the association of CD3zeta did not require the GPI anchor of TSA-1. Finally, tyrosine phosphorylation of CD3zeta chains was induced after stimulation with anti-TSA-1, suggesting that a functional association between these two molecules also exists. These results imply that the physical association to CD3zeta underlies a regulatory role of TSA-1/Sca-2 in the TCR-signaling pathway.     

Heat shock cognate protein 71-associated peptides function as an epitope for Toxoplasma gondii-specific CD4+ CTL

HLA-DR-restricted CD4+ cytotoxic T-lymphocyte (CTL) lines specific for Toxoplasma gondii (T. gondii)-infected melanoma cells have been established from peripheral blood lymphocytes (PBLs) of a patient with chronic toxoplasmosis. The role of heat shock cognate protein (HSC) 71 in antigen (Ag) processing and presentation of T. gondii-infected melanoma cells to these CD4+ CTL lines was investigated. A human melanoma cell line (P36) pulsed with T. gondii-infected P36 cell-derived HSC71 was lysed by a T. gondii-specific CD4+ CTL line (Tx-HSC-1). The Tx-HSC-1 also killed T. gondii-infected P36 cells. The lytic activity of Tx-HSC-1 against P36 cells pulsed with T. gondii-infected P36 cell-derived HSC71 was inhibited by monoclonal antibodies (mAbs) against HSC71. Anti-human leukocyte antigen (HLA)-DR mAb also partially blocked the lytic activity, whereas anti-HLA-A,B,C mAb did not block the lytic activity. In addition, a flow cytometric analysis with these specific mAbs against HSC71 showed HSC71 to be expressed on the cell surface of T. gondii-infected P36 cells as well as uninfected P36 cells. These data indicate that HSC71 molecules are expressed on human melanoma cell line P36, and that HSC71 may play a potential role in Ag presentation and processing of T. gondii-infected P36 cells to CD4+ CTL.     

Simultaneous induction of multiple antigen-specific cytotoxic T lymphocytes in nonhuman primates by immunization with a mixture of four Plasmodium falciparum DNA plasmids

CD8(+) T cells have been implicated as critical effector cells in protective immunity against malaria parasites developing within hepatocytes. A vaccine that protects against malaria by inducing CD8(+) T cells will probably have to include multiple epitopes on the same protein or different proteins, because of parasite polymorphism and genetic restriction of T-cell responses. To determine if CD8(+) T-cell responses against multiple P. falciparum proteins can be induced in primates by immunization with plasmid DNA, rhesus monkeys were immunized intramuscularly with a mixture of DNA plasmids encoding four P. falciparum proteins or with individual plasmids. All six monkeys immunized with PfCSP DNA, seven of nine immunized with PfSSP2 DNA, and five of six immunized with PfExp-1 or PfLSA-1 DNA had detectable antigen-specific cytotoxic T lymphocytes (CTL) after in vitro restimulation of peripheral blood mononuclear cells. CTL activity was genetically restricted and dependent on CD8(+) T cells. By providing the first evidence for primates that immunization with a mixture of DNA plasmids induces CD8(+) T-cell responses against all the components of the mixture, these studies provide the foundation for multigene immunization of humans.     

Is epsilon-aminocaproic acid as effective as aprotinin in reducing bleeding with cardiac surgery?: a meta-analysis

Infection of BALB/c mice with Trypanosoma cruzi resulted in up-regulated expression of Fas and Fas ligand (FasL) mRNA by splenic CD4+ T cells, activation-induced CD4+ T cell death (AICD), and in Fas: FasL-mediated cytotoxicity. When CD4+ T cells from infected mice were co-cultured with T. cruzi-infected macrophages, onset of AICD exacerbated parasite replication. CD4+ T cells from T. cruzi-infected FasL-deficient BALB gld/gld mice had no detectable AICD in vitro and their activation with anti-TCR did not exacerbate T. cruzi replication in macrophages. However, infection of BALB gld/gld mice with T. cruzi resulted in higher and more prolonged parasitemia, compared to wild-type mice. Secretion of Th2 cytokines IL-10 and IL-4 by CD4+ T cells from infected gld mice was markedly increased, compared to controls. In addition, in vivo injection of anti-IL-4 mAb, but not of an isotype control mAb, reduced parasitemia in both gld and wild-type mice. These results indicate that, besides controlling CD4+ T cell AICD and parasite replication in vitro, an intact Fas: FasL pathway also controls the host cytokine response to T. cruzi infection in vivo, being required to prevent an exacerbated Th2-type immune response to the parasite.     

Teratogenic effects and distribution of cadmium (Cd2+) administered via osmotic minipumps to gravid CF-1 mice

Cytotoxic T lymphocytes (CTL) specific against autologous human cervical cancer cells were generated in vitro from peripheral blood leukocytes (PBL) from four patients with non-keratinized epidermoid carcinoma. For this purpose, these patients' PBL were co-cultured for 28 days either with IL-2 or a mixture of IL-2, IFN-gamma and TNF-alpha in the presence of autologous tumour cells (ATC). Our results showed that these CTL were highly cytotoxic for ATC, weakly cytotoxic for heterologous cervical cancer tumour cells, and not cytotoxic for carcinoma cell lines, normal cervix cells nor autologous PBL. Proliferation and cytotoxicity against ATC were greater when the PBL were activated with the three cytokines. These CTL had a CD4:CD8 ratio of 1:1, were CD16- and CD45RO+ and their killing activity was inhibited by antibodies against CD3, CD8 and MHC-class I but not by antibodies against CD4, CD16 or HLA-class II. The possibility of generating specific CTL in long term cultures for cervical cancer therapy is also discussed.     

Fas ligand-mediated lysis of self bystander targets by human papillomavirus-specific CD8+ cytotoxic T lymphocytes

Mouse cytotoxic T lymphocytes (CTL) reactive with a H-2Db-presented 9-mer peptide of the human papillomavirus type 16 protein E7(49-57) (RAHYNIVTF) were generated from the spleen cells of wild-type C57BL/6 (B6) or B6 perforin-deficient (B6.P0) mice. CD8(+) B6 CTL displayed peptide-specific perforin- and Fas-mediated lysis of E7-transfected mouse RMA lymphoma cells (RMA-E7), while CD8(+) CTL from B6.P0 mice lysed RMA-E7 cells via Fas ligand (FasL) exclusively. Rapid and efficient lysis of syngeneic bystander B6 blasts or RMA cells by either B6 or B6.P0 Ag-activated CTL was mediated by a FasL-Fas mechanism. Fas-resistant bystanders were not lysed, nor were allogeneic Fas-sensitive C3H/HeJ (H-2(k)) or BALB/c (H-2(d)) bystander blasts. Interestingly, however, phorbol myristate acetate-ionomycin preactivation of B6.P0 effectors enabled lysis of allogeneic H-2(k) and H-2(d) bystanders even in the absence of antigenic stimulation. Lysis of syngeneic bystander cells was always FasL-Fas dependent and required effector-bystander contact and, in particular, an interaction between CTL LFA-1 and bystander ICAM-1. Thus, in the context of major histocompatibility complex class I molecule-peptide ligation of the T-cell receptors of CD8(+) CTL, neighboring bystander cells that are syngeneic and Fas sensitive and express the adhesion molecule ICAM-1 are potential targets of CTL attack.     

Trypanosoma cruzi: protective response of vaccinated mice is mediated by CD8+ cells, prevents signs of polyclonal T lymphocyte activation, and allows restoration of a resting immune state after challenge

Currently, there is no vaccine available against Chagas' disease. Immune abnormalities induced by T. cruzi pose particular difficulties for vaccine development, since immunological memory must be able to overcome them to prevent spread of infection/sequelae. We have previously demonstrated that experimental vaccination with live CL-14 trypomastigotes does not induce polyclonal lymphocyte activation, immunosuppression, or pathology and efficiently immunizes against virulent T. cruzi. Herein we show that: (1) expansion of CD4+ and CD8+ subsets peaks 2 weeks after infective challenge in both challenged-vaccinated mice and infected controls, but the former exhibit a smaller increase in blastogenesis and in the numbers of activated CD11a(hi)CD4+ and CD11a(hi)CD8+ cells; (2) in long-term-vaccinated mice, expansion of activated subsets (CD62Llo/- and CD11a(hi)) is accelerated among CD8+ PBL 1 week after challenge; (3) challenged-vaccinated mice retract the CD8+-activated subset 5 weeks after challenge, different from infected controls; (4) protection conferred by CL-14 immunization can be adoptively transferred to na?ve recipients with lymphocyte suspensions, and prior depletion of CD8+ (but not of CD4+) cells abolishes protective immunity. Our findings indicate that protective immunity generated by CL-14 immunization involves a transient CD8+ recall response and is capable of preventing the signs of polyclonal lymphocyte activation induced by virulent challenge.     

Protection of mice against Trypanosoma cruzi by immunization with paraflagellar rod proteins requires T cell, but not B cell, function

Previous studies have shown that immunization of mice with the paraflagellar rod proteins (PAR) of Trypanosoma cruzi induces an immune response capable of protecting mice against an otherwise lethal challenge with this parasite. Herein, we define immunologic responses that do or do not play a critical role in PAR-mediated protection. Firstly, PAR-immunized Ab-deficient (muMT) strain mice survived an otherwise lethal T. cruzi challenge, indicating that a B cell response is not required for PAR-induced immunity. However, beta2m -/- mice, which are severely deficient in MHC class I and TCR alphabeta+ CD8+ CD4- T cells, did not survive challenge infection following PAR immunization, indicating that MHC class I/CD8+ T cell function is necessary for protection induced by PAR immunization. Surprisingly, PAR-immunized mice depleted of CD4+ T cells survived a T. cruzi challenge for >84 days postinfection while maintaining a parasitemia that is generally thought to be lethal (i.e., >10(6) trypomastigotes/ml), thus associating CD4+ T cell function with the process of parasite clearance. Consistent with this association, CD4+ T cells from PAR-immunized mice released INF-gamma and stimulated T. cruzi-infected macrophages to release nitric oxide. The importance of IFN-gamma in PAR-induced protective immunity is further indicated by the observation that PAR-immunized INF-gamma knockout mice developed an extremely high parasitemia and did not survive a challenge infection. Thus, while Ab-mediated immune mechanisms are not required for protection induced by PAR immunization, T cell responses are necessary for both elimination of bloodstream parasites and survival.     

Cryopreservation of teeth. Organizational aspects of a tissue bank for tooth tissues

Squamous cell carcinoma (SCC) is one of the most common cancers in human. SCC, particularly, esophageal and lung SCC are relatively resistant to currently available regimens of chemotherapy or radiation therapy. Therefore, development of a specific immunotherapy using tumor specific cytotoxic T lymphocytes (CTL) would be important to offer other treatment modalities. However, generation of HLA class I-restricted CTL recognizing SCC has been rarely reported. We established the HLA A2601-restricted CTL cell line recognizing a peptide antigen expressed on SCC. This CD4- CD8+ cytotoxic T lymphocyte (KE-4 CTL) cell line was established in a patient with esophageal cancer. The KE-4 CTL recognized a peptide antigen on esohageal and lung SCC in an HLA A2601-restricted manner as evaluated by cytotoxity against a panel of tumor cells, transfection experiments with HLA A2601 cDNA, and reconstitution with eleted peptides. None of normal cells tested was lysed by this CTL. These results suggest the exstence of HLA A2601-restricted CTL precursors recognizing a peptide antigen on SCC in a patient with esophageal cancer.     

Histocompatibility leukocyte antigen-A2-restricted and tumor-specific cytotoxic T lymphocytes from tumor-infiltrating lymphocytes of patient with testicular embryonal cancer

T lymphocytes play an important role in tumor rejection. To understand T cell-mediated specific immunity at the tumor site of testicular embryonal cancer, we investigated whether interleukin-2 (IL-2)-activated tumor-infiltrating lymphocytes (TIL) of a patient with testicular embryonal cancer show histocompatibility leukocyte antigen (HLA)-class I-restricted and tumor-specific cytotoxicity. We established a CD3+CD4-CD8+ cytotoxic T lymphocyte (CTL) line from the IL-2-activated TIL of a 37-year-old patient with testicular embryonal cancer. A 6 h 51Cr-release assay was performed to measure the cytotoxicity of the CTL. The CD3+CD4-CD8+ CTL line showed cytotoxicity against HLA-A2+ tumor cells, including freshly isolated autologous tumor cells, adenocarcinoma cell lines from various organs (lung, breast, pancreas, colon and kidney) and squamous cell carcinomas (esophagus and oral cavity). No other cell lines examined, including an autologous tumor cell line and HLA-A2- tumor cell lines, were lysed by this CTL line. These results suggest the existence of HLA-A2-restricted and tumor-specific CTL at the tumor site of testicular embryonal cancer.     

Dengue virus protein recognition by virus-specific murine CD8+ cytotoxic T lymphocytes

The identification of the protein targets for dengue virus-specific T lymphocytes may be useful for planning the development of subunit vaccines against dengue. We studied the recognition by murine dengue virus-specific major histocompatibility complex class I-restricted, CD8+ cytotoxic T lymphocytes (CTL) of dengue virus proteins using recombinant vaccinia viruses containing segments of the dengue virus genome. CTL from H-2k mice recognized a single serotype-cross-reactive epitope on the nonstructural (NS) protein NS3. CTL from H-2b mice recognized a serotype-cross-reactive epitope that was localized to NS4a or NS4b. CTL from H-2d mice recognized at least three epitopes: a serotype-specific epitope on one of the structural proteins, a serotype-cross-reactive epitope on NS3, and a serotype-cross-reactive epitope on NS1 or NS2a. Our findings demonstrate the limited recognition of dengue virus proteins by CTL from three inbred mouse strains and the predominance of CTL epitopes on dengue virus nonstructural proteins, particularly NS3. Since human dengue virus-specific CTL show similar patterns of recognition, these findings suggest that nonstructural proteins should be considered in designing vaccines against dengue.     

Cell population changes during atrophy and regeneration of rat parotid gland

Melan-A/MART-1 is a melanoma differentiation antigen that is recognized by a high proportion of cytolytic T lymphocyte (CTL) clones derived from human leukocyte antigen (HLA)-A2+ melanoma patients. Whereas peptide Melan-A/ MART-1(27-35) was originally defined as the immunodominant CTL epitope, we have previously reported that peptide Melan-A/MART-1(26-35) was recognized more efficiently by the majority of tumor-reactive CTL clones. As demonstrated here, CTL populations generated from blood lymphocytes of either melanoma patients or healthy individuals after in vitro stimulation with peptide Melan-A/MART-1(26-35) killed specifically HLA-A2+ Melan-A+ allogeneic melanoma cells, thus suggesting their potential use in adoptive immunotherapy. We characterized the surface phenotype of the circulating CTL precursors (CTLp), which respond to in vitro stimulation with peptide Melan-A/MART-1(26-35). In melanoma patients, these CTLp predominantly expressed the CD45RO memory marker. In contrast, they were mainly, although not exclusively, found in the CD45RA subpopulation of CD8 T cells in healthy individuals. The demonstration that Melan-A/MART-1-specific CTLp in peripheral blood lymphocytes from HLA-A2+ patients with metastatic melanoma express a memory phenotype provides direct evidence that in vivo priming of this antigen may occur during tumor progression.     

Expression of the CD8alpha beta-heterodimer on CD8(+) T lymphocytes in peripheral blood lymphocytes of human immunodeficiency virus- and human immunodeficiency virus+ individuals

CD8(+) T lymphocytes play a pivotal role in controlling human immunodeficiency virus (HIV)-1 replication in vivo. We have performed four-color flow cytometric analysis of CD8(+) peripheral blood lymphocytes (PBL) from 21 HIV-1 seronegative and 103 seropositive individuals to explore the phenotypic heterogeneity of CD8beta-chain expression on CD8(+) T lymphocytes and to clarify how its expression on CD8(+) T lymphocytes may relate to acquired immunodeficiency syndrome (AIDS) clinical progression. We showed that the single monoclonal antibody (MoAb) 2ST8-5H7, directed against the CD8alpha beta-heterodimer, identifies CD8(+) T lymphocytes as effectively as the conventional combination of anti-CD3 and anti-CD8alpha antibodies. However, we detected a significantly lower mean fluorescence (MF) of anti-CD8alpha beta staining on PBL from HIV-1 seropositive donors as compared with seronegative donors. In fact, CD8(+) T lymphocytes from HIV-1-infected individuals with the lowest CD4 counts showed the lowest levels of CD8alpha beta MF. To explore further this change in CD8alpha beta expression, we assessed the expression of 14 different cell surface molecules on CD8alpha beta+ T lymphocytes of PBL from 11 HIV-1 seronegative and 22 HIV-1 seropositive individuals. The MF of anti-CD8alpha beta staining was significantly reduced on CD8(+) T lymphocyte subsets that showed immunophenotypic evidence of activation. The subset of lymphocytes expressing low levels of CD8alpha beta expressed higher levels of activation, adhesion, and cytotoxic-associated molecules and was predominantly CD45RO+ and CD28(-). Finally, we monitored the expression of the CD8alpha beta-heterodimer on PBL of eight HIV-1-infected individuals over a 16-week period after the initiation of highly active antiretroviral therapy (HAART), including zidovudine (ZDV), lamivudine (3TC), and indinavir (IDV), and found a significant increase in the expression of the CD8alpha beta-heterodimer. These results suggest that antibodies recognizing the CD8alpha beta-heterodimer are useful tools to specifically identify CD8(+) T lymphocytes. Moreover, the quantitative monitoring of CD8alpha beta expression allows the detection of discrete CD8(+) T lymphocyte subsets and may be useful for assessing the immune status of individuals infected with HIV-1.     

Tumor regression in mice following vaccination with human papillomavirus E7 recombinant protein in PROVAX

Induction of CD8+ cytotoxic T lymphocytes (CTLs) specific for human papillomavirus (HPV) antigens provides an attractive strategy for immunotherapy of HPV-related cancers in humans. In this study, we investigated the potential of utilizing soluble E7 protein of HPV 16 in an adjuvant formulation, PROVAX as a vaccine against a progressively growing E7 transfected K1735-X21 (H-2k) metastatic melanoma cells (HOPE2) in a mouse model. Vaccination of HOPE2 tumor bearing mice (C3H) with E7 protein in PROVAX resulted in significant inhibition of tumor growth, compared to mice vaccinated with E7 in Alum or saline. In vivo depletion of CD8+ or CD4+ cells indicated that CD8+ cells are the major effector cells in mediating the anti-tumor activity in this model. Furthermore, E7-specific CTL activity in vitro was detected in tumor bearing mice vaccinated with E7-PROVAX. Our studies suggest that recombinant HPV antigens in combination with PROVAX could serve as an effective subunit vaccine to stimulate tumor specific CD8+ T cell mediated immunity against HPV-related cancers.     

Liver damage preferentially results from CD8(+) T cells triggered by high affinity peptide antigens

Little is understood of the anatomical fate of activated T lymphocytes and the consequences they have on the tissues into which they migrate. Previous work has suggested that damaged lymphocytes migrate to the liver. This study compares class I versus class II major histocompatibility complex (MHC)-restricted ovalbumin-specific T cell antigen receptor (TCR) transgenic mice to demonstrate that after in vivo activation with antigen the emergence of CD4(-)CD8(-)B220(+) T cells occurs more frequently from a CD8(+) precursor than from CD4(+) T cells. Furthermore, this change in phenotype is conferred only by the high affinity native peptide antigen and not by lower affinity peptide variants. After activation of CD8(+) cells with only the high affinity peptide, there is also a dramatically increased number of liver lymphocytes with accompanying extensive hepatocyte damage and elevation of serum aspartate transaminase. This was not observed in mice bearing a class II MHC-restricted TCR. The findings show that CD4(-)CD8(-)B220(+) T cells preferentially derive from a CD8(+) precursor after a high intensity TCR signal. After activation, T cells can migrate to the liver and induce hepatocyte damage, and thereby serve as a model of autoimmune hepatitis.     

The theory and practice of health education applied to nursing: a bi-polar approach

Because CD1-restricted T cells lack CD4 but produce IFN-gamma in response to nonpeptide mycobacterial antigens, they could play a unique role in immunity to tuberculosis. We studied CD1-restricted T cells in the context of HIV infection by expanding CD4(-) T cell lines from 10 HIV-infected patients. Upon stimulation with Mycobacterium tuberculosis antigen or upon exposure to macrophages infected with M. tuberculosis, these T cell lines proliferated, produced IFN-gamma, and showed cytolytic T cell (CTL) activity against macrophages pulsed with mycobacterial antigen, findings consistent with a protective role against M. tuberculosis. Anti-CD1b antibodies abrogated T cell proliferation, IFN-gamma production, and CTL activity, demonstrating that these T cells are CD1 restricted. IFN-gamma production in response to M. tuberculosis was enhanced by antitransforming growth factor-beta in 8/10 lines, and by IL-15 in 2/10 lines. IFN-gamma production was augmented in a nonantigen-specific manner by IL-12 in 4/8 lines. When live HIV was cocultured with CD1-restricted T cell lines, p24 antigen and proviral DNA were not detected, indicating that the T cells were not infectable with HIV. Vaccination strategies aimed at activation and expansion of M. tuberculosis-reactive CD1-restricted T cells in HIV-infected patients may constitute a novel means to provide protection against tuberculosis, while minimizing the risk of enhancing HIV replication through stimulation of CD4(+) cells.     

Immunization with a single major histocompatibility complex class I-restricted cytotoxic T-lymphocyte recognition epitope of herpes simplex virus type 2 confers protective immunity

We have evaluated the potential of conferring protective immunity to herpes simplex virus type 2 (HSV-2) by selectively inducing an HSV-specific CD8(+) cytotoxic T-lymphocyte (CTL) response directed against a single major histocompatibility complex class I-restricted CTL recognition epitope. We generated a recombinant vaccinia virus (rVV-ES-gB498-505) which expresses the H-2Kb-restricted, HSV-1/2-cross-reactive CTL recognition epitope, HSV glycoprotein B residues 498 to 505 (SSIEFARL) (gB498-505), fused to the adenovirus type 5 E3/19K endoplasmic reticulum insertion sequence (ES). Mucosal immunization of C57BL/6 mice with this recombinant vaccinia virus induced both a primary CTL response in the draining lymph nodes and a splenic memory CTL response directed against HSV gB498-505. To determine the ability of the gB498-505-specific memory CTL response to provide protection from HSV infection, immunized mice were challenged with a lethal dose of HSV-2 strain 186 by the intranasal (i.n.) route. Development of the gB498-505-specific CTL response conferred resistance in 60 to 75% of mice challenged with a lethal dose of HSV-2 and significantly reduced the levels of infectious virus in the brains and trigeminal ganglia of challenged mice. Finally, i.n. immunization of C57BL/6 mice with either a recombinant influenza virus or a recombinant vaccinia virus expressing HSV gB498-505 without the ES was also demonstrated to induce an HSV-specific CTL response and provide protection from HSV infection. This finding confirms that the induction of an HSV-specific CTL response directed against a single epitope is sufficient for conferring protective immunity to HSV. Our findings support the role of CD8(+) T cells in the control of HSV infection of the central nervous system and suggest the potential importance of eliciting HSV-specific mucosal CD8(+) CTL in HSV vaccine design.