Adoptive Immunotherapy against Experimental Visceral Leishmaniasis with CD8+ T Cells Requires the Presence of Cognate Antigen
http://www.100md.com
感染与免疫杂志 2006年第1期
Department of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, Keppel Street, London, WC1E 7HT United Kingdom
Wellcome Trust Laboratories for Molecular Parasitology, Department of Biological Sciences, Imperial College London, London, SW7 2AZ United Kingdom
ABSTRACT
CD8+ T cells have a protective role in experimental visceral leishmaniasis. However, the observation that inflammatory cytokines induce bystander activation of CD8+ T cells questions the need for antigen-dependent effector function. Here, we demonstrate that successful adoptive immunotherapy with CD8+ T cells is strictly dependent upon the presence of cognate antigen.
TEXT
CD8+ T cells provide a major component of host defense against a variety of viral and nonviral pathogens and have been implicated as a major component of vaccine-induced immunity against some of the most important human pathogens (1-3). CD8+ T cells have been demonstrated to play a role in various experimental models of leishmaniasis, including infection with Leishmania major (2, 13), L. mexicana (10), L. infantum (21), and L. donovani (8, 14, 19, 20). However, the study of CD8+ T-cell function in leishmaniasis has been hampered compared to the study of their role in viral and bacterial infections by the lack of suitable tools for directly measuring the function of Leishmania-specific CD8+ T cells and for discriminating between antigen-dependent and potentially antigen-independent effector functions (10). In two models of cutaneous leishmaniasis, antigen-specific CD8+ T cells generated to defined Leishmania-expressed antigens have failed either to exert protection in vivo (22) or to kill infected macrophages in vitro (10). Recent studies demonstrating that a variety of inflammatory cytokines can mediate antigen-independent "bystander activation" of CD8+ memory and effector T cells, leading to host protection in different disease models (3, 6, 9), now question whether the effector function of CD8+ T cells in experimental leishmaniasis requires the presence of cognate antigen. Here, we have used adoptive CD8+ T-cell immunotherapy during experimental visceral leishmaniasis to directly examine this question.
We recently described a vaccination model in which host protection against L. donovani is mediated at least in part by CD8+ T cells with specificity for an amastigote-expressed antigen, hydrophilic acylated surface protein B1 (HASPB1) (18). HASPB1 is unusual in being targeted for expression at the plasma membrane by a unique N terminus which specifies its dual acylation (4), and this property may facilitate phagosomal presentation for class I processing (5). We therefore made use of this property of HASPB1 and generated transgenic L. donovani expressing a HASP1-18-OVA fusion protein. Full details of the generation of these and other Leishmania lines expressing HASP1-18-OVA will be published elsewhere. Briefly, the sequence encoding the N-terminal 18 amino acids of the HASPs (4) was cloned in frame with the OVA open reading frame and inserted into the pSSU vector for gene targeting by homologous recombination into a ribosomal DNA locus (11). Log-phase promastigotes of L. donovani (LV9) were electroporated with 40 μg of PacI/PmeI-digested DNA, and the resulting transfectants were maintained in 40 μg/ml hygromycin (Invitrogen) prior to single-cell cloning. Clones were screened for integration of the linear targeting fragment by Southern blot analysis of genomic DNA and for OVA expression by immunoblotting analysis of parasite lysate (Fig. 1). These transgenic parasites are formally designated Ld18SrRNA::HASP1-18-OVA (herein referred to as PINK). In order to recover infectivity, positive PINK clones were transformed to amastigotes (16) and maintained by serial passage in hamsters. Expression of OVA was maintained over several months of passage in the absence of drug selection. Semiquantitative analysis of the levels of OVA expression indicated that these transgenic parasites expressed approximately 2 ng OVA/106 promastigotes or approximately 0.05% of the total amount of protein, similar to the expression level of HASPB in Leishmania (4) and somewhat higher than the level of OVA expressed in Leishmania from the episomal vector pX (7). Clones of transgenic OVA-expressing L. donovani were fully infective to mice, and host immunity was similarly organ specific, as has been described for wild-type infections. Clone 6, with maximal expression of OVA as determined by immunoblotting, was chosen for in vivo studies.
To determine whether adoptive immunotherapy with CD8+ T cells altered the course of experimental visceral leishmaniasis and to define whether such an effect required in vivo antigen recognition, B6.CD45.1 mice were infected intravenously with 2 x 107 wild-type L. donovani (LV9) amastigotes or transgenic PINK amastigotes. At day 21 postinfection, adoptive transfer of 1 x 106 T-cell receptor transgenic OVA-specific OT-I CD8+ T cells was performed (see below). Hepatic and splenic parasite burdens were determined by examining methanol-fixed, Giemsa-stained tissue impression smears 7 days after transfer (24). All animal procedures were approved by the LSHTM Animal Procedures Ethics Committee and the United Kingdom Home Office.
In the first series of experiments, we wished to evaluate whether in vitro-generated cytotoxic CD8+ T cells were host protective. OT-I cytotoxic T lymphocytes (CTL) were generated by incubation of nave OT-I cells with 1 nM OVA257-264, 350 ng/ml recombinant interleukin 4 (rIL-4), and 10 ng/ml rIL-2 for 3 days, followed by a further 2 days with cytokine and no peptide (1). CTL activity was confirmed by lysis of EG7-OVA (12) but not parental EL4 cells (data not shown). As shown in Fig. 2, the transfer of OT-I CTL into mice infected with wild-type L. donovani had no impact on parasite growth in either the liver or the spleen. In contrast, OT-I cells exerted significant control over the course of infection with PINK amastigotes in both target tissues. In the liver, this was evident as leishmanicidal activity, whereas in the spleen, the effect was restricted to being leishmanistatic. Together, these data both indicate a requirement for cognate antigen for successful immunotherapy by CTL and, by implication, demonstrate that the inflammatory environment created by wild-type L. donovani infection is insufficient to induce bystander effector responses from these CTL.
In the second series of experiments, we confirmed and extended these findings to other populations of CD8+ T cells. CD8+ memory T cells have been divided into two subsets. Effector memory T cells continually recirculate through peripheral tissues, whereas central memory T cells, by virtue of their high levels of expression of CD62L and CCR7, are endowed with lymph node homing capacity (17). CD8+ T cells resembling effector and central memory cells were derived in vitro as described previously (23). Briefly, splenocytes from nave OT-1 transgenic mice were incubated with 10 μg/ml OVA257-264 for 1 h at 37°C, washed, and cultured for a further 48 h. Cells were then washed and incubated for a further 9 days with either 20 ng/ml rIL-2 (effector memory-like OT-IIL-2) or 20 ng/ml rIL-15 (central memory-like OT-IIL-15). Cytokines were changed every second day. OT-IIL-2 and OT-IIL-15 cells used for transfer were >88% CD62Llo and >75% CD62Lhi, respectively. In some experiments, cells were sorted on the basis of CD62L to >99% purity, with similar results.
As with OT-I CTL, neither OT-IIL-2 nor OT-IIL-15 cells had any effect on the course of wild-type L. donovani infection (Fig. 3A). In contrast, the transfer of OT-IIL-2 cells had a rapid impact on the course of PINK infection (Fig. 3B). Hepatic parasite burden 48 h after transfer was reduced by 61% ± 6% compared to levels in control mice (P < 0.001), an effect mirrored in the spleen (59% ± 5%; P < 0.001). This therapeutic effect was sustained and/or improved when determined 7 days after transfer (54% ± 6% and 84% ± 6% reduction in liver and spleen, respectively). In comparison, OT-IIL-15 and OT-I cells adoptively transferred directly after isolation from nave OT-I transgenic mice did not exert a statistically significant effect on host resistance until day 7 after transfer, though at this time they were as effective as OT-IIL-2 cells (Fig. 3B). Collectively, these data indicate that cells with memory cell characteristics, as well as nave OVA-specific CD8+ T cells, are capable of transferring significant protection against ongoing infection, albeit with differing kinetics of action. As with transferred CTL, we found no evidence for bystander activation of host-protective function. Rather, therapeutic benefit was strictly dependent upon infection with parasites expressing the cognate antigen recognized by these CD8+ T cells.
In summary, we have addressed a basic yet fundamental issue related to CD8+ T-cell function in the later stages of experimental visceral leishmaniasis, namely, whether cognate antigen is required for CD8+ T cells to exert host-protective function. Our data indicate that this is indeed the case and suggest that future interventions based on increasing antigen recognition by endogenous CD8+ T cells during infection may be beneficial. Additional studies are required to confirm why bystander activation of effector function in CD8+ T cells does not occur to any significant extent in this model, in spite of extensive inflammation and evidence for bystander proliferation of both CD4+ (15) and CD8+ (15a) T cells. Furthermore, the model system that we describe here should also be useful for analyzing other facets of CD8+ T-cell biology, including the differential requirements for antigen-dependent activation of nave versus effector/memory CD8+ T cells and the mechanisms underlying their protective effects.
ACKNOWLEDGMENTS
We thank the staff of the Biological Services Facility for excellent animal husbandry and T. Aebischer for the kind provision of pSSU-int.
This work was supported by grants from the British Medical Research Council and the Wellcome Trust. R.P. was a recipient of an MRC Training Fellowship. S.P. was a recipient of an Imperial College Department of Biochemistry Postgraduate Bursary.
These authors contributed equally to this work.
Present address: Lymphocyte Biology Section, Laboratory of Immunology, NIAID/National Institutes of Health, Bethesda, MD 20892.
Present address: Division of Biomedical Sciences, Johns Hopkins in Singapore, 31 Biopolis Way, 02-01 The Nanos, Singapore 138669, Republic of Singapore.
|| Present address: Division of Infection and Immunity, Walter and Eliza Hall Institute, 1G Royal Parade, Parkville, Victoria 3050, Australia.
Present address: Immunology and Infection Unit, Dept. of Biology, University of York, P.O. Box 373, York, YO10 5YW United Kingdom.
REFERENCES
1. Bathe, O. F., N. Dalyot-Herman, and T. R. Malek. 2001. IL-2 during in vitro priming promotes subsequent engraftment and successful adoptive tumor immunotherapy by persistent memory phenotypic CD8(+) T cells. J. Immunol. 167:4511-4517.
2. Belkaid, Y., E. Von Stebut, S. Mendez, R. Lira, E. Caler, S. Bertholet, M. C. Udey, and D. Sacks. 2002. CD8+ T cells are required for primary immunity in C57BL/6 mice following low-dose, intradermal challenge with Leishmania major. J. Immunol. 168:3992-4000.
3. Berg, R. E., E. Crossley, S. Murray, and J. Forman. 2003. Memory CD8+ T cells provide innate immune protection against Listeria monocytogenes in the absence of cognate antigen. J. Exp. Med. 198:1583-1593.
4. Denny, P. W., S. Gokool, D. G. Russell, M. C. Field, and D. F. Smith. 2000. Acylation-dependent protein export in Leishmania. J. Biol. Chem. 275:11017-11025.
5. Houde, M., S. Bertholet, E. Gagnon, S. Brunet, G. Goyette, A. Laplante, M. F. Princiotta, P. Thibault, D. Sacks, and M. Desjardins. 2003. Phagosomes are competent organelles for antigen cross-presentation. Nature 425:402-406.
6. Kamath, A. T., C. E. Sheasby, and D. F. Tough. 2005. Dendritic cells and NK cells stimulate bystander T cell activation in response to TLR agonists through secretion of IFN-alpha beta and IFN-gamma. J. Immunol. 174:767-776.
7. Kaye, P. M., C. Coburn, M. McCrossan, and S. M. Beverley. 1993. Antigens targeted to the Leishmania phagolysosome are processed for CD4+ T cell recognition. Eur. J. Immunol. 23:2311-2319.
8. Kaye, P. M., A. Cooke, T. Lund, M. Wattie, and J. M. Blackwell. 1992. Altered course of visceral leishmaniasis in mice expressing transgenic I-E molecules. Eur. J. Immunol. 22:357-364.
9. Lertmemongkolchai, G., G. Cai, C. A. Hunter, and G. J. Bancroft. 2001. Bystander activation of CD8+ T cells contributes to the rapid production of IFN-gamma in response to bacterial pathogens. J. Immunol. 166:1097- 1105.
10. Lopez, J. A., J. H. LeBowitz, S. M. Beverley, H. G. Rammensee, and P. Overath. 1993. Leishmania mexicana promastigotes induce cytotoxic T lymphocytes in vivo that do not recognize infected macrophages. Eur. J. Immunol. 23:217-223.
11. Misslitz, A., J. C. Mottram, P. Overath, and T. Aebischer. 2000. Targeted integration into a rRNA locus results in uniform and high level expression of transgenes in Leishmania amastigotes. Mol. Biochem. Parasitol. 107:251-261.
12. Moore, M. W., F. R. Carbone, and M. J. Bevan. 1988. Introduction of soluble protein into the class I pathway of antigen processing and presentation. Cell 54:777-785.
13. Muller, I., P. Kropf, J. A. Louis, and G. Milon. 1994. Expansion of gamma interferon-producing CD8+ T cells following secondary infection of mice immune to Leishmania major. Infect. Immun. 62:2575-2581.
14. Murray, H. W. 2005. Prevention of relapse after chemotherapy in a chronic intracellular infection: mechanisms in experimental visceral leishmaniasis. J. Immunol. 174:4916-4923.
15. Polley, R., S. Zubairi, and P. M. Kaye. 2005. The fate of heterologous CD4+ T cells during Leishmania donovani infection. Eur. J. Immunol. 35:498-504.
15. Polley, R., S. L. Sanos, S. Prickett, A. Hague, and P. M. Kaye. 2005. Chronic Leishmania donovani infection promotes bystander CD8+-T-cell expansion and heterologous immunity. Infect. Immun. 73:7996-8001.
16. Saar, Y., A. Ransford, E. Waldman, S. Mazareb, S. Amin-Spector, J. Plumblee, S. J. Turco, and D. Zilberstein. 1998. Characterization of developmentally-regulated activities in axenic amastigotes of Leishmania donovani. Mol. Biochem. Parasitol. 95:9-20.
17. Sallusto, F., D. Lenig, R. Forster, M. Lipp, and A. Lanzavecchia. 1999. Two subsets of memory T lymphocytes with distinct homing potentials and effector functions. Nature 401:708-712.
18. Stager, S., J. Alexander, A. C. Kirby, M. Botto, N. V. Rooijen, D. F. Smith, F. Brombacher, and P. M. Kaye. 2003. Natural antibodies and complement are endogenous adjuvants for vaccine-induced CD8+ T-cell responses. Nat. Med. 9:1287-1292.
19. Stern, J. J., M. J. Oca, B. Y. Rubin, S. L. Anderson, and H. W. Murray. 1988. Role of L3T4+ and LyT-2+ cells in experimental visceral leishmaniasis. J. Immunol. 140:3971-3977.
20. Tsagozis, P., E. Karagouni, and E. Dotsika. 2003. CD8(+) T cells with parasite-specific cytotoxic activity and a Tc1 profile of cytokine and chemokine secretion develop in experimental visceral leishmaniasis. Parasite Immunol. 25:569-579.
21. Tsagozis, P., E. Karagouni, and E. Dotsika. 2005. Function of CD8+ T lymphocytes in a self-curing mouse model of visceral leishmaniasis. Parasitol. Int. 54:139-146.
22. Wang, Z. E., S. L. Reiner, F. Hatam, F. P. Heinzel, J. Bouvier, C. W. Turck, and R. M. Locksley. 1993. Targeted activation of CD8 cells and infection of beta 2-microglobulin-deficient mice fail to confirm a primary protective role for CD8 cells in experimental leishmaniasis. J. Immunol. 151:2077-2086.
23. Weninger, W., M. A. Crowley, N. Manjunath, and U. H. von Andrian. 2001. Migratory properties of naive, effector, and memory CD8(+) T cells. J. Exp. Med. 194:953-966.
24. Zubairi, S., S. L. Sanos, S. Hill, and P. M. Kaye. 2004. Immunotherapy with OX40L-Fc or anti-CTLA-4 enhances local tissue responses and killing of Leishmania donovani. Eur. J. Immunol. 34:1433-1440.(Rosalind Polley , Simona )
Wellcome Trust Laboratories for Molecular Parasitology, Department of Biological Sciences, Imperial College London, London, SW7 2AZ United Kingdom
ABSTRACT
CD8+ T cells have a protective role in experimental visceral leishmaniasis. However, the observation that inflammatory cytokines induce bystander activation of CD8+ T cells questions the need for antigen-dependent effector function. Here, we demonstrate that successful adoptive immunotherapy with CD8+ T cells is strictly dependent upon the presence of cognate antigen.
TEXT
CD8+ T cells provide a major component of host defense against a variety of viral and nonviral pathogens and have been implicated as a major component of vaccine-induced immunity against some of the most important human pathogens (1-3). CD8+ T cells have been demonstrated to play a role in various experimental models of leishmaniasis, including infection with Leishmania major (2, 13), L. mexicana (10), L. infantum (21), and L. donovani (8, 14, 19, 20). However, the study of CD8+ T-cell function in leishmaniasis has been hampered compared to the study of their role in viral and bacterial infections by the lack of suitable tools for directly measuring the function of Leishmania-specific CD8+ T cells and for discriminating between antigen-dependent and potentially antigen-independent effector functions (10). In two models of cutaneous leishmaniasis, antigen-specific CD8+ T cells generated to defined Leishmania-expressed antigens have failed either to exert protection in vivo (22) or to kill infected macrophages in vitro (10). Recent studies demonstrating that a variety of inflammatory cytokines can mediate antigen-independent "bystander activation" of CD8+ memory and effector T cells, leading to host protection in different disease models (3, 6, 9), now question whether the effector function of CD8+ T cells in experimental leishmaniasis requires the presence of cognate antigen. Here, we have used adoptive CD8+ T-cell immunotherapy during experimental visceral leishmaniasis to directly examine this question.
We recently described a vaccination model in which host protection against L. donovani is mediated at least in part by CD8+ T cells with specificity for an amastigote-expressed antigen, hydrophilic acylated surface protein B1 (HASPB1) (18). HASPB1 is unusual in being targeted for expression at the plasma membrane by a unique N terminus which specifies its dual acylation (4), and this property may facilitate phagosomal presentation for class I processing (5). We therefore made use of this property of HASPB1 and generated transgenic L. donovani expressing a HASP1-18-OVA fusion protein. Full details of the generation of these and other Leishmania lines expressing HASP1-18-OVA will be published elsewhere. Briefly, the sequence encoding the N-terminal 18 amino acids of the HASPs (4) was cloned in frame with the OVA open reading frame and inserted into the pSSU vector for gene targeting by homologous recombination into a ribosomal DNA locus (11). Log-phase promastigotes of L. donovani (LV9) were electroporated with 40 μg of PacI/PmeI-digested DNA, and the resulting transfectants were maintained in 40 μg/ml hygromycin (Invitrogen) prior to single-cell cloning. Clones were screened for integration of the linear targeting fragment by Southern blot analysis of genomic DNA and for OVA expression by immunoblotting analysis of parasite lysate (Fig. 1). These transgenic parasites are formally designated Ld18SrRNA::HASP1-18-OVA (herein referred to as PINK). In order to recover infectivity, positive PINK clones were transformed to amastigotes (16) and maintained by serial passage in hamsters. Expression of OVA was maintained over several months of passage in the absence of drug selection. Semiquantitative analysis of the levels of OVA expression indicated that these transgenic parasites expressed approximately 2 ng OVA/106 promastigotes or approximately 0.05% of the total amount of protein, similar to the expression level of HASPB in Leishmania (4) and somewhat higher than the level of OVA expressed in Leishmania from the episomal vector pX (7). Clones of transgenic OVA-expressing L. donovani were fully infective to mice, and host immunity was similarly organ specific, as has been described for wild-type infections. Clone 6, with maximal expression of OVA as determined by immunoblotting, was chosen for in vivo studies.
To determine whether adoptive immunotherapy with CD8+ T cells altered the course of experimental visceral leishmaniasis and to define whether such an effect required in vivo antigen recognition, B6.CD45.1 mice were infected intravenously with 2 x 107 wild-type L. donovani (LV9) amastigotes or transgenic PINK amastigotes. At day 21 postinfection, adoptive transfer of 1 x 106 T-cell receptor transgenic OVA-specific OT-I CD8+ T cells was performed (see below). Hepatic and splenic parasite burdens were determined by examining methanol-fixed, Giemsa-stained tissue impression smears 7 days after transfer (24). All animal procedures were approved by the LSHTM Animal Procedures Ethics Committee and the United Kingdom Home Office.
In the first series of experiments, we wished to evaluate whether in vitro-generated cytotoxic CD8+ T cells were host protective. OT-I cytotoxic T lymphocytes (CTL) were generated by incubation of nave OT-I cells with 1 nM OVA257-264, 350 ng/ml recombinant interleukin 4 (rIL-4), and 10 ng/ml rIL-2 for 3 days, followed by a further 2 days with cytokine and no peptide (1). CTL activity was confirmed by lysis of EG7-OVA (12) but not parental EL4 cells (data not shown). As shown in Fig. 2, the transfer of OT-I CTL into mice infected with wild-type L. donovani had no impact on parasite growth in either the liver or the spleen. In contrast, OT-I cells exerted significant control over the course of infection with PINK amastigotes in both target tissues. In the liver, this was evident as leishmanicidal activity, whereas in the spleen, the effect was restricted to being leishmanistatic. Together, these data both indicate a requirement for cognate antigen for successful immunotherapy by CTL and, by implication, demonstrate that the inflammatory environment created by wild-type L. donovani infection is insufficient to induce bystander effector responses from these CTL.
In the second series of experiments, we confirmed and extended these findings to other populations of CD8+ T cells. CD8+ memory T cells have been divided into two subsets. Effector memory T cells continually recirculate through peripheral tissues, whereas central memory T cells, by virtue of their high levels of expression of CD62L and CCR7, are endowed with lymph node homing capacity (17). CD8+ T cells resembling effector and central memory cells were derived in vitro as described previously (23). Briefly, splenocytes from nave OT-1 transgenic mice were incubated with 10 μg/ml OVA257-264 for 1 h at 37°C, washed, and cultured for a further 48 h. Cells were then washed and incubated for a further 9 days with either 20 ng/ml rIL-2 (effector memory-like OT-IIL-2) or 20 ng/ml rIL-15 (central memory-like OT-IIL-15). Cytokines were changed every second day. OT-IIL-2 and OT-IIL-15 cells used for transfer were >88% CD62Llo and >75% CD62Lhi, respectively. In some experiments, cells were sorted on the basis of CD62L to >99% purity, with similar results.
As with OT-I CTL, neither OT-IIL-2 nor OT-IIL-15 cells had any effect on the course of wild-type L. donovani infection (Fig. 3A). In contrast, the transfer of OT-IIL-2 cells had a rapid impact on the course of PINK infection (Fig. 3B). Hepatic parasite burden 48 h after transfer was reduced by 61% ± 6% compared to levels in control mice (P < 0.001), an effect mirrored in the spleen (59% ± 5%; P < 0.001). This therapeutic effect was sustained and/or improved when determined 7 days after transfer (54% ± 6% and 84% ± 6% reduction in liver and spleen, respectively). In comparison, OT-IIL-15 and OT-I cells adoptively transferred directly after isolation from nave OT-I transgenic mice did not exert a statistically significant effect on host resistance until day 7 after transfer, though at this time they were as effective as OT-IIL-2 cells (Fig. 3B). Collectively, these data indicate that cells with memory cell characteristics, as well as nave OVA-specific CD8+ T cells, are capable of transferring significant protection against ongoing infection, albeit with differing kinetics of action. As with transferred CTL, we found no evidence for bystander activation of host-protective function. Rather, therapeutic benefit was strictly dependent upon infection with parasites expressing the cognate antigen recognized by these CD8+ T cells.
In summary, we have addressed a basic yet fundamental issue related to CD8+ T-cell function in the later stages of experimental visceral leishmaniasis, namely, whether cognate antigen is required for CD8+ T cells to exert host-protective function. Our data indicate that this is indeed the case and suggest that future interventions based on increasing antigen recognition by endogenous CD8+ T cells during infection may be beneficial. Additional studies are required to confirm why bystander activation of effector function in CD8+ T cells does not occur to any significant extent in this model, in spite of extensive inflammation and evidence for bystander proliferation of both CD4+ (15) and CD8+ (15a) T cells. Furthermore, the model system that we describe here should also be useful for analyzing other facets of CD8+ T-cell biology, including the differential requirements for antigen-dependent activation of nave versus effector/memory CD8+ T cells and the mechanisms underlying their protective effects.
ACKNOWLEDGMENTS
We thank the staff of the Biological Services Facility for excellent animal husbandry and T. Aebischer for the kind provision of pSSU-int.
This work was supported by grants from the British Medical Research Council and the Wellcome Trust. R.P. was a recipient of an MRC Training Fellowship. S.P. was a recipient of an Imperial College Department of Biochemistry Postgraduate Bursary.
These authors contributed equally to this work.
Present address: Lymphocyte Biology Section, Laboratory of Immunology, NIAID/National Institutes of Health, Bethesda, MD 20892.
Present address: Division of Biomedical Sciences, Johns Hopkins in Singapore, 31 Biopolis Way, 02-01 The Nanos, Singapore 138669, Republic of Singapore.
|| Present address: Division of Infection and Immunity, Walter and Eliza Hall Institute, 1G Royal Parade, Parkville, Victoria 3050, Australia.
Present address: Immunology and Infection Unit, Dept. of Biology, University of York, P.O. Box 373, York, YO10 5YW United Kingdom.
REFERENCES
1. Bathe, O. F., N. Dalyot-Herman, and T. R. Malek. 2001. IL-2 during in vitro priming promotes subsequent engraftment and successful adoptive tumor immunotherapy by persistent memory phenotypic CD8(+) T cells. J. Immunol. 167:4511-4517.
2. Belkaid, Y., E. Von Stebut, S. Mendez, R. Lira, E. Caler, S. Bertholet, M. C. Udey, and D. Sacks. 2002. CD8+ T cells are required for primary immunity in C57BL/6 mice following low-dose, intradermal challenge with Leishmania major. J. Immunol. 168:3992-4000.
3. Berg, R. E., E. Crossley, S. Murray, and J. Forman. 2003. Memory CD8+ T cells provide innate immune protection against Listeria monocytogenes in the absence of cognate antigen. J. Exp. Med. 198:1583-1593.
4. Denny, P. W., S. Gokool, D. G. Russell, M. C. Field, and D. F. Smith. 2000. Acylation-dependent protein export in Leishmania. J. Biol. Chem. 275:11017-11025.
5. Houde, M., S. Bertholet, E. Gagnon, S. Brunet, G. Goyette, A. Laplante, M. F. Princiotta, P. Thibault, D. Sacks, and M. Desjardins. 2003. Phagosomes are competent organelles for antigen cross-presentation. Nature 425:402-406.
6. Kamath, A. T., C. E. Sheasby, and D. F. Tough. 2005. Dendritic cells and NK cells stimulate bystander T cell activation in response to TLR agonists through secretion of IFN-alpha beta and IFN-gamma. J. Immunol. 174:767-776.
7. Kaye, P. M., C. Coburn, M. McCrossan, and S. M. Beverley. 1993. Antigens targeted to the Leishmania phagolysosome are processed for CD4+ T cell recognition. Eur. J. Immunol. 23:2311-2319.
8. Kaye, P. M., A. Cooke, T. Lund, M. Wattie, and J. M. Blackwell. 1992. Altered course of visceral leishmaniasis in mice expressing transgenic I-E molecules. Eur. J. Immunol. 22:357-364.
9. Lertmemongkolchai, G., G. Cai, C. A. Hunter, and G. J. Bancroft. 2001. Bystander activation of CD8+ T cells contributes to the rapid production of IFN-gamma in response to bacterial pathogens. J. Immunol. 166:1097- 1105.
10. Lopez, J. A., J. H. LeBowitz, S. M. Beverley, H. G. Rammensee, and P. Overath. 1993. Leishmania mexicana promastigotes induce cytotoxic T lymphocytes in vivo that do not recognize infected macrophages. Eur. J. Immunol. 23:217-223.
11. Misslitz, A., J. C. Mottram, P. Overath, and T. Aebischer. 2000. Targeted integration into a rRNA locus results in uniform and high level expression of transgenes in Leishmania amastigotes. Mol. Biochem. Parasitol. 107:251-261.
12. Moore, M. W., F. R. Carbone, and M. J. Bevan. 1988. Introduction of soluble protein into the class I pathway of antigen processing and presentation. Cell 54:777-785.
13. Muller, I., P. Kropf, J. A. Louis, and G. Milon. 1994. Expansion of gamma interferon-producing CD8+ T cells following secondary infection of mice immune to Leishmania major. Infect. Immun. 62:2575-2581.
14. Murray, H. W. 2005. Prevention of relapse after chemotherapy in a chronic intracellular infection: mechanisms in experimental visceral leishmaniasis. J. Immunol. 174:4916-4923.
15. Polley, R., S. Zubairi, and P. M. Kaye. 2005. The fate of heterologous CD4+ T cells during Leishmania donovani infection. Eur. J. Immunol. 35:498-504.
15. Polley, R., S. L. Sanos, S. Prickett, A. Hague, and P. M. Kaye. 2005. Chronic Leishmania donovani infection promotes bystander CD8+-T-cell expansion and heterologous immunity. Infect. Immun. 73:7996-8001.
16. Saar, Y., A. Ransford, E. Waldman, S. Mazareb, S. Amin-Spector, J. Plumblee, S. J. Turco, and D. Zilberstein. 1998. Characterization of developmentally-regulated activities in axenic amastigotes of Leishmania donovani. Mol. Biochem. Parasitol. 95:9-20.
17. Sallusto, F., D. Lenig, R. Forster, M. Lipp, and A. Lanzavecchia. 1999. Two subsets of memory T lymphocytes with distinct homing potentials and effector functions. Nature 401:708-712.
18. Stager, S., J. Alexander, A. C. Kirby, M. Botto, N. V. Rooijen, D. F. Smith, F. Brombacher, and P. M. Kaye. 2003. Natural antibodies and complement are endogenous adjuvants for vaccine-induced CD8+ T-cell responses. Nat. Med. 9:1287-1292.
19. Stern, J. J., M. J. Oca, B. Y. Rubin, S. L. Anderson, and H. W. Murray. 1988. Role of L3T4+ and LyT-2+ cells in experimental visceral leishmaniasis. J. Immunol. 140:3971-3977.
20. Tsagozis, P., E. Karagouni, and E. Dotsika. 2003. CD8(+) T cells with parasite-specific cytotoxic activity and a Tc1 profile of cytokine and chemokine secretion develop in experimental visceral leishmaniasis. Parasite Immunol. 25:569-579.
21. Tsagozis, P., E. Karagouni, and E. Dotsika. 2005. Function of CD8+ T lymphocytes in a self-curing mouse model of visceral leishmaniasis. Parasitol. Int. 54:139-146.
22. Wang, Z. E., S. L. Reiner, F. Hatam, F. P. Heinzel, J. Bouvier, C. W. Turck, and R. M. Locksley. 1993. Targeted activation of CD8 cells and infection of beta 2-microglobulin-deficient mice fail to confirm a primary protective role for CD8 cells in experimental leishmaniasis. J. Immunol. 151:2077-2086.
23. Weninger, W., M. A. Crowley, N. Manjunath, and U. H. von Andrian. 2001. Migratory properties of naive, effector, and memory CD8(+) T cells. J. Exp. Med. 194:953-966.
24. Zubairi, S., S. L. Sanos, S. Hill, and P. M. Kaye. 2004. Immunotherapy with OX40L-Fc or anti-CTLA-4 enhances local tissue responses and killing of Leishmania donovani. Eur. J. Immunol. 34:1433-1440.(Rosalind Polley , Simona )