Complex Memory T-Cell Phenotypes Revealed by Coexp
http://www.100md.com
病菌学杂志 2005年第6期
Institute for Medical Microbiology and Hygiene, Department of Immunology, University of Freiburg, Freiburg, Germany
ABSTRACT
Antigen-experienced T cells have been divided into CD62L+ CCR7+ central memory (TCM) and CD62L– CCR7– effector memory (TEM) cells. Here, we examined coexpression of CD62L and CCR7 in lymphocytic choriomeningitis virus-specific memory CD8 T cells from both lymphoid and nonlymphoid tissues. Three main points emerged: firstly, memory cells frequently expressed a mixed CD62L– CCR7+ phenotype that differed from the phenotypes of classical TEM and TCM cells; secondly, TCM cells were not restricted to lymphoid organs but were also present in significant numbers in nonlymphoid tissues; and thirdly, a major shift from a TCM to TEM phenotype was found in memory cells that had been stimulated repetitively with antigen.
TEXT
Based on expression of CD62L and CCR7, memory T cells have been divided into two main subsets (14): central memory T cells (TCM), expressing both CD62L and CCR7 and representing nonpolarized antigen-experienced cells, and effector memory cells (TEM), lacking CD62L/CCR7 cell surface expression but capable of performing immediate effector cell functions. Studies with mice further demonstrated that in response to antigen, a significant number of T cells leave secondary lymphoid tissues and reside as long-lived memory cells in nonlymphoid tissues (8, 10-12). Memory T cells isolated from nonlymphoid tissues were found to exhibit cytolytic activities and to produce inflammatory cytokines, in contrast to memory cells from secondary lymphoid organs (11, 12). The results of these mouse models fit nicely into the TCM/TEM concept (14), which was based primarily on phenotypic and functional analysis of human memory T cells in vitro. However, the in vivo CD62L/CCR7 expression patterns of true antigen-specific memory T cells isolated from different lymphoid and nonlymphoid tissues have not yet been characterized in detail.
To address this issue, we used a well-characterized adoptive transfer system with P14 T-cell receptor transgenic cells specific for the major histocompatibility complex class I-restricted GP33 epitope of lymphocytic choriomeningitis virus (LCMV) to generate bona fide memory CD8 T cells in vivo (18). Briefly, 105 Thy1.1+ P14 T cells were transferred intravenously into C57BL/6 (B6) mice, followed by infection with 200 PFU of LCMV-WE. LCMV infection in this transfer model leads to high viral titers (106 to 107 PFU/g) in the spleen on day 4 after infection. By day 8 postinfection, LCMV is cleared almost completely (<102 PFU/g of tissue) in all organs (19). Tissue distribution and cell surface phenotype of P14 memory cells were determined 7 weeks after LCMV infection by flow cytometry using antibodies specific for Thy1.1 (clone OX-7) and CD62L (clone Mel14) that had been purchased from BD Pharmingen (San Diego, Calif.). Cell surface expression of CCR7 was determined by a chimeric CCL19-immunoglobulin fusion protein (15). Lymphocytes from lymphoid organs and from perfused liver and lungs were isolated and stained by using standard protocols (13, 15). Collagenase treatment was omitted, since it resulted in partial digestion of CD62L during lymphocyte isolation from liver but not from lung, spleen, and lymph nodes (data not shown). Cells were analyzed on a FACSCalibur flow cytometer (BD Biosciences). Similar to observations in other viral systems (8, 10, 11), higher frequencies of LCMV-specific P14 memory cells were found in liver and lung than in blood, spleen, and lymph nodes (Fig. 1A).
Examination of P14 memory cells for coexpression of CD62L and CCR7 revealed three major memory T-cell subsets: CD62L+ CCR7+, CD62L– CCR7+, and CD62L– CCR7– cells (Fig. 1B). According to previous definitions (14), CD62L+ CCR7+ and CD62L– CCR7– cells were classified as P14 TCM and P14 TEM cells, respectively. The third memory cell population expressing CCR7 in the absence of CD62L was not previously noted and is therefore defined operationally in this study as intermediate memory T cells (TIM). Analysis of P14 memory cells from lymphoid and nonlymphoid tissues revealed the following picture (Fig. 1C): P14 TCM (38% ± 13%), P14 TEM (28 ± 11%), and P14 TIM (25 ± 9%) memory cell populations were present in roughly equal numbers in the blood, P14 TCM cells (60% ± 10%) predominated in the spleen, and P14 TCM (55% ± 9%) and P14 TIM (30% ± 13%) cells represented the two major cell populations in inguinal lymph nodes. In the lung, most of the P14 cells exhibited a TEM phenotype (51% ± 9%), followed by TCM (26% ± 7%) and TIM (16% ± 6%).
The differentiation pathway of TCM and TEM cells is a matter of debate. Sallusto et al. (14) proposed that antigen stimulation leads to TCM cells that can then further differentiate into TEM, whereas data obtained by Wherry et al. (16) with the LCMV mouse model favored the opposite view. The authors of the latter study proposed a conversion of TEM to TCM cells over time after viral clearance. Differentiation of memory T cells may also be influenced by repeated antigenic stimulation (1). To mimic a priming and boosting regimen in our transfer model, memory P14 T cells, isolated from the spleen of LCMV immune mice 7 weeks after infection, were retransferred into B6 mice followed by a second LCMV infection (Fig. 2A). This resulted in a strong wave of proliferation of P14 memory cells followed by a decline similar to that observed in primary transfers with naive P14 cells (Fig. 2B). Interestingly, the CD62L/CCR7 phenotype of these secondary P14 memory cells, also isolated 7 weeks postinfection, differed considerably from primary P14 memory T cells. In all organs analyzed, a clear shift from P14 TCM to P14 TEM cells was apparent (Fig. 2C). This was particularly evident in the blood, where the fraction of P14 TCM cells dropped from 38% ± 13% to 8 ± 4% and the proportion of P14 TEM cells rose from 28% ± 11% to 56% ± 12%. To provide evidence that secondary P14 TEM cells were indeed derived from P14 TCM cells, 105 CD62L+ CCR7+ P14 T cells from primary transfers (5 weeks postinfection) were purified by cell sorting and were retransferred into B6 recipient mice, followed by LCMV infection. The CD62L/CCR7 phenotypes, analyzed 8 weeks after transfer and infection (Fig. 3), were similar to the data shown in Fig. 2C obtained with nonpurified P14 memory cells. Comparison of secondary versus primary P14 memory cells again revealed a significant shift from a TCM to a TEM phenotype in all organs analyzed. Hence, our data indicate that repetitive antigen stimulation induces a differentiation from P14 TCM to P14 TEM cells in vivo.
In conclusion, our data demonstrate that memory CD8 T cells frequently express a mixed CD62L– CCR7+ phenotype that differs from phenotypes of classical CD62L+ CCR7+ TCM and CD62L– CCR7– TEM cells. In other words, expression of CD62L and CCR7 in bona fide memory CD8 T cells in vivo overlaps only partially. This point is important, since some groups use expression of CD62L alone to distinguish TCM and TEM cells (5, 16, 17), whereas others use expression of CCR7 (2, 4, 9, 14, 15) for the same purpose. Very recently, coexpression of CD62L and CCR7 in CD4 T cells from influenza virus-infected mice has been described (3). Similar to our data with CD8 T cells, CD4 T cells carrying a mixed CD62L– CCR7+ phenotype were also found. However, in contrast to our approach, bona fide antigen-specific memory T cells could not be identified in the latter study. Another important aspect of our report concerns the presence of sizeable numbers of P14 memory T cells in liver and lung that expressed a TCM phenotype. Thus, despite being equipped with two major lymph node homing receptors, CD62L (7) and CCR7 (6), these cells were located in nonlymphoid organs. This result cautions against the exclusive use of cell surface phenotypes to predict tissue localization. Finally, the shift from a TCM to a TEM phenotype in P14 memory cells that had been stimulated twice with antigen indicates that TCM/TEM differentiation pathways could be influenced by a priming/boosting regimen. This result is also relevant for vaccine strategies, since it predicts that repetitive antigen exposure favors TEM differentiation.
ACKNOWLEDGMENTS
We thank S. Batsford for comments on the manuscript.
This work was supported by the Deutsche Forschungsgemeinschaft (PI 295/5).
REFERENCES
Appay, V., P. R. Dunbar, M. Callan, P. Klenerman, G. M. Gillespie, L. Papagno, G. S. Ogg, A. King, F. Lechner, C. A. Spina, S. Little, D. V. Havlir, D. D. Richman, N. Gruener, G. Pape, A. Waters, P. Easterbrook, M. Salio, V. Cerundolo, A. J. McMichael, and S. L. Rowland-Jones. 2002. Memory CD8+ T cells vary in differentiation phenotype in different persistent virus infections. Nat. Med. 8:379-385.
Champagne, P., G. S. Ogg, A. S. King, C. Knabenhans, K. Ellefsen, M. Nobile, V. Appay, G. P. Rizzardi, S. Fleury, M. Lipp, R. Forster, S. Rowland-Jones, R. P. Sekaly, A. J. McMichael, and G. Pantaleo. 2001. Skewed maturation of memory HIV-specific CD8 T lymphocytes. Nature 410:106-111.
Debes, G. F., K. Bonhagen, T. Wolff, U. Kretschmer, S. Krautwald, T. Kamradt, and A. Hamann. 2004. CC chemokine receptor 7 expression by effector/memory CD4+ T cells depends on antigen specificity and tissue localization during influenza A virus infection. J. Virol. 78:7528-7535.
Debes, G. F., U. E. Hopken, and A. Hamann. 2002. In vivo differentiated cytokine-producing CD4+ T cells express functional CCR7. J. Immunol. 168:5441-5447.
Ely, K. H., A. D. Roberts, and D. L. Woodland. 2003. Cutting edge: effector memory CD8+ T cells in the lung airways retain the potential to mediate recall responses. J. Immunol. 171:3338-3342.
Forster, R., A. Schubel, D. Breitfeld, E. Kremmer, I. Renner-Muller, E. Wolf, and M. Lipp. 1999. CCR7 coordinates the primary immune response by establishing functional microenvironments in secondary lymphoid organs. Cell 99:23-33.
Gallatin, W. M., I. L. Weissman, and E. C. Butcher. 1983. A cell-surface molecule involved in organ-specific homing of lymphocytes. Nature 304:30-34.
Hogan, R. J., L. S. Cauley, K. H. Ely, T. Cookenham, A. D. Roberts, J. W. Brennan, S. Monard, and D. L. Woodland. 2002. Long-term maintenance of virus-specific effector memory CD8+ T cells in the lung airways depends on proliferation. J. Immunol. 169:4976-4981.
Kim, C. H., L. Rott, E. J. Kunkel, M. C. Genovese, D. P. Andrew, L. Wu, and E. C. Butcher. 2001. Rules of chemokine receptor association with T cell polarization in vivo. J. Clin. Investig. 108:1331-1339.
Marshall, D. R., S. J. Turner, G. T. Belz, S. Wingo, S. Andreansky, M. Y. Sangster, J. M. Riberdy, T. Liu, M. Tan, and P. C. Doherty. 2001. Measuring the diaspora for virus-specific CD8+ T cells. Proc. Natl. Acad. Sci. USA 98:6313-6318.
Masopust, D., V. Vezys, A. L. Marzo, and L. Lefrancois. 2001. Preferential localization of effector memory cells in nonlymphoid tissue. Science 291:2413-2417.
Reinhardt, R. L., A. Khoruts, R. Merica, T. Zell, and M. K. Jenkins. 2001. Visualizing the generation of memory CD4 T cells in the whole body. Nature 410:101-105.
Roth, E., and H. Pircher. 2004. IFN-gamma promotes Fas ligand- and perforin-mediated liver cell destruction by cytotoxic CD8 T cells. J. Immunol. 172:1588-1594.
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.
Unsoeld, H., S. Krautwald, D. Voehringer, U. Kunzendorf, and H. Pircher. 2002. Cutting edge: CCR7+ and CCR7– memory T cells do not differ in immediate effector cell function. J. Immunol. 169:638-641.
Wherry, E. J., V. Teichgraber, T. C. Becker, D. Masopust, S. M. Kaech, R. Antia, U. H. von Andrian, and R. Ahmed. 2003. Lineage relationship and protective immunity of memory CD8 T cell subsets. Nat. Immunol. 4:225-234.
Wolint, P., M. R. Betts, R. A. Koup, and A. Oxenius. 2004. Immediate cytotoxicity but not degranulation distinguishes effector and memory subsets of CD8+ T cells. J. Exp. Med. 199:925-936.
Zimmerman, C., K. Brduscha-Riem, C. Blaser, R. M. Zinkernagel, and H. Pircher. 1996. Visualization, characterization, and turnover of CD8+ memory T cells in virus-infected hosts. J. Exp. Med. 183:1367-1375.
Zimmermann, C., A. Prevost-Blondel, C. Blaser, and H. Pircher. 1999. Kinetics of the response of naive and memory CD8 T cells to antigen: similarities and differences. Eur. J. Immunol. 29:284-290.(Heike Unsoeld and Hanspet)
ABSTRACT
Antigen-experienced T cells have been divided into CD62L+ CCR7+ central memory (TCM) and CD62L– CCR7– effector memory (TEM) cells. Here, we examined coexpression of CD62L and CCR7 in lymphocytic choriomeningitis virus-specific memory CD8 T cells from both lymphoid and nonlymphoid tissues. Three main points emerged: firstly, memory cells frequently expressed a mixed CD62L– CCR7+ phenotype that differed from the phenotypes of classical TEM and TCM cells; secondly, TCM cells were not restricted to lymphoid organs but were also present in significant numbers in nonlymphoid tissues; and thirdly, a major shift from a TCM to TEM phenotype was found in memory cells that had been stimulated repetitively with antigen.
TEXT
Based on expression of CD62L and CCR7, memory T cells have been divided into two main subsets (14): central memory T cells (TCM), expressing both CD62L and CCR7 and representing nonpolarized antigen-experienced cells, and effector memory cells (TEM), lacking CD62L/CCR7 cell surface expression but capable of performing immediate effector cell functions. Studies with mice further demonstrated that in response to antigen, a significant number of T cells leave secondary lymphoid tissues and reside as long-lived memory cells in nonlymphoid tissues (8, 10-12). Memory T cells isolated from nonlymphoid tissues were found to exhibit cytolytic activities and to produce inflammatory cytokines, in contrast to memory cells from secondary lymphoid organs (11, 12). The results of these mouse models fit nicely into the TCM/TEM concept (14), which was based primarily on phenotypic and functional analysis of human memory T cells in vitro. However, the in vivo CD62L/CCR7 expression patterns of true antigen-specific memory T cells isolated from different lymphoid and nonlymphoid tissues have not yet been characterized in detail.
To address this issue, we used a well-characterized adoptive transfer system with P14 T-cell receptor transgenic cells specific for the major histocompatibility complex class I-restricted GP33 epitope of lymphocytic choriomeningitis virus (LCMV) to generate bona fide memory CD8 T cells in vivo (18). Briefly, 105 Thy1.1+ P14 T cells were transferred intravenously into C57BL/6 (B6) mice, followed by infection with 200 PFU of LCMV-WE. LCMV infection in this transfer model leads to high viral titers (106 to 107 PFU/g) in the spleen on day 4 after infection. By day 8 postinfection, LCMV is cleared almost completely (<102 PFU/g of tissue) in all organs (19). Tissue distribution and cell surface phenotype of P14 memory cells were determined 7 weeks after LCMV infection by flow cytometry using antibodies specific for Thy1.1 (clone OX-7) and CD62L (clone Mel14) that had been purchased from BD Pharmingen (San Diego, Calif.). Cell surface expression of CCR7 was determined by a chimeric CCL19-immunoglobulin fusion protein (15). Lymphocytes from lymphoid organs and from perfused liver and lungs were isolated and stained by using standard protocols (13, 15). Collagenase treatment was omitted, since it resulted in partial digestion of CD62L during lymphocyte isolation from liver but not from lung, spleen, and lymph nodes (data not shown). Cells were analyzed on a FACSCalibur flow cytometer (BD Biosciences). Similar to observations in other viral systems (8, 10, 11), higher frequencies of LCMV-specific P14 memory cells were found in liver and lung than in blood, spleen, and lymph nodes (Fig. 1A).
Examination of P14 memory cells for coexpression of CD62L and CCR7 revealed three major memory T-cell subsets: CD62L+ CCR7+, CD62L– CCR7+, and CD62L– CCR7– cells (Fig. 1B). According to previous definitions (14), CD62L+ CCR7+ and CD62L– CCR7– cells were classified as P14 TCM and P14 TEM cells, respectively. The third memory cell population expressing CCR7 in the absence of CD62L was not previously noted and is therefore defined operationally in this study as intermediate memory T cells (TIM). Analysis of P14 memory cells from lymphoid and nonlymphoid tissues revealed the following picture (Fig. 1C): P14 TCM (38% ± 13%), P14 TEM (28 ± 11%), and P14 TIM (25 ± 9%) memory cell populations were present in roughly equal numbers in the blood, P14 TCM cells (60% ± 10%) predominated in the spleen, and P14 TCM (55% ± 9%) and P14 TIM (30% ± 13%) cells represented the two major cell populations in inguinal lymph nodes. In the lung, most of the P14 cells exhibited a TEM phenotype (51% ± 9%), followed by TCM (26% ± 7%) and TIM (16% ± 6%).
The differentiation pathway of TCM and TEM cells is a matter of debate. Sallusto et al. (14) proposed that antigen stimulation leads to TCM cells that can then further differentiate into TEM, whereas data obtained by Wherry et al. (16) with the LCMV mouse model favored the opposite view. The authors of the latter study proposed a conversion of TEM to TCM cells over time after viral clearance. Differentiation of memory T cells may also be influenced by repeated antigenic stimulation (1). To mimic a priming and boosting regimen in our transfer model, memory P14 T cells, isolated from the spleen of LCMV immune mice 7 weeks after infection, were retransferred into B6 mice followed by a second LCMV infection (Fig. 2A). This resulted in a strong wave of proliferation of P14 memory cells followed by a decline similar to that observed in primary transfers with naive P14 cells (Fig. 2B). Interestingly, the CD62L/CCR7 phenotype of these secondary P14 memory cells, also isolated 7 weeks postinfection, differed considerably from primary P14 memory T cells. In all organs analyzed, a clear shift from P14 TCM to P14 TEM cells was apparent (Fig. 2C). This was particularly evident in the blood, where the fraction of P14 TCM cells dropped from 38% ± 13% to 8 ± 4% and the proportion of P14 TEM cells rose from 28% ± 11% to 56% ± 12%. To provide evidence that secondary P14 TEM cells were indeed derived from P14 TCM cells, 105 CD62L+ CCR7+ P14 T cells from primary transfers (5 weeks postinfection) were purified by cell sorting and were retransferred into B6 recipient mice, followed by LCMV infection. The CD62L/CCR7 phenotypes, analyzed 8 weeks after transfer and infection (Fig. 3), were similar to the data shown in Fig. 2C obtained with nonpurified P14 memory cells. Comparison of secondary versus primary P14 memory cells again revealed a significant shift from a TCM to a TEM phenotype in all organs analyzed. Hence, our data indicate that repetitive antigen stimulation induces a differentiation from P14 TCM to P14 TEM cells in vivo.
In conclusion, our data demonstrate that memory CD8 T cells frequently express a mixed CD62L– CCR7+ phenotype that differs from phenotypes of classical CD62L+ CCR7+ TCM and CD62L– CCR7– TEM cells. In other words, expression of CD62L and CCR7 in bona fide memory CD8 T cells in vivo overlaps only partially. This point is important, since some groups use expression of CD62L alone to distinguish TCM and TEM cells (5, 16, 17), whereas others use expression of CCR7 (2, 4, 9, 14, 15) for the same purpose. Very recently, coexpression of CD62L and CCR7 in CD4 T cells from influenza virus-infected mice has been described (3). Similar to our data with CD8 T cells, CD4 T cells carrying a mixed CD62L– CCR7+ phenotype were also found. However, in contrast to our approach, bona fide antigen-specific memory T cells could not be identified in the latter study. Another important aspect of our report concerns the presence of sizeable numbers of P14 memory T cells in liver and lung that expressed a TCM phenotype. Thus, despite being equipped with two major lymph node homing receptors, CD62L (7) and CCR7 (6), these cells were located in nonlymphoid organs. This result cautions against the exclusive use of cell surface phenotypes to predict tissue localization. Finally, the shift from a TCM to a TEM phenotype in P14 memory cells that had been stimulated twice with antigen indicates that TCM/TEM differentiation pathways could be influenced by a priming/boosting regimen. This result is also relevant for vaccine strategies, since it predicts that repetitive antigen exposure favors TEM differentiation.
ACKNOWLEDGMENTS
We thank S. Batsford for comments on the manuscript.
This work was supported by the Deutsche Forschungsgemeinschaft (PI 295/5).
REFERENCES
Appay, V., P. R. Dunbar, M. Callan, P. Klenerman, G. M. Gillespie, L. Papagno, G. S. Ogg, A. King, F. Lechner, C. A. Spina, S. Little, D. V. Havlir, D. D. Richman, N. Gruener, G. Pape, A. Waters, P. Easterbrook, M. Salio, V. Cerundolo, A. J. McMichael, and S. L. Rowland-Jones. 2002. Memory CD8+ T cells vary in differentiation phenotype in different persistent virus infections. Nat. Med. 8:379-385.
Champagne, P., G. S. Ogg, A. S. King, C. Knabenhans, K. Ellefsen, M. Nobile, V. Appay, G. P. Rizzardi, S. Fleury, M. Lipp, R. Forster, S. Rowland-Jones, R. P. Sekaly, A. J. McMichael, and G. Pantaleo. 2001. Skewed maturation of memory HIV-specific CD8 T lymphocytes. Nature 410:106-111.
Debes, G. F., K. Bonhagen, T. Wolff, U. Kretschmer, S. Krautwald, T. Kamradt, and A. Hamann. 2004. CC chemokine receptor 7 expression by effector/memory CD4+ T cells depends on antigen specificity and tissue localization during influenza A virus infection. J. Virol. 78:7528-7535.
Debes, G. F., U. E. Hopken, and A. Hamann. 2002. In vivo differentiated cytokine-producing CD4+ T cells express functional CCR7. J. Immunol. 168:5441-5447.
Ely, K. H., A. D. Roberts, and D. L. Woodland. 2003. Cutting edge: effector memory CD8+ T cells in the lung airways retain the potential to mediate recall responses. J. Immunol. 171:3338-3342.
Forster, R., A. Schubel, D. Breitfeld, E. Kremmer, I. Renner-Muller, E. Wolf, and M. Lipp. 1999. CCR7 coordinates the primary immune response by establishing functional microenvironments in secondary lymphoid organs. Cell 99:23-33.
Gallatin, W. M., I. L. Weissman, and E. C. Butcher. 1983. A cell-surface molecule involved in organ-specific homing of lymphocytes. Nature 304:30-34.
Hogan, R. J., L. S. Cauley, K. H. Ely, T. Cookenham, A. D. Roberts, J. W. Brennan, S. Monard, and D. L. Woodland. 2002. Long-term maintenance of virus-specific effector memory CD8+ T cells in the lung airways depends on proliferation. J. Immunol. 169:4976-4981.
Kim, C. H., L. Rott, E. J. Kunkel, M. C. Genovese, D. P. Andrew, L. Wu, and E. C. Butcher. 2001. Rules of chemokine receptor association with T cell polarization in vivo. J. Clin. Investig. 108:1331-1339.
Marshall, D. R., S. J. Turner, G. T. Belz, S. Wingo, S. Andreansky, M. Y. Sangster, J. M. Riberdy, T. Liu, M. Tan, and P. C. Doherty. 2001. Measuring the diaspora for virus-specific CD8+ T cells. Proc. Natl. Acad. Sci. USA 98:6313-6318.
Masopust, D., V. Vezys, A. L. Marzo, and L. Lefrancois. 2001. Preferential localization of effector memory cells in nonlymphoid tissue. Science 291:2413-2417.
Reinhardt, R. L., A. Khoruts, R. Merica, T. Zell, and M. K. Jenkins. 2001. Visualizing the generation of memory CD4 T cells in the whole body. Nature 410:101-105.
Roth, E., and H. Pircher. 2004. IFN-gamma promotes Fas ligand- and perforin-mediated liver cell destruction by cytotoxic CD8 T cells. J. Immunol. 172:1588-1594.
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.
Unsoeld, H., S. Krautwald, D. Voehringer, U. Kunzendorf, and H. Pircher. 2002. Cutting edge: CCR7+ and CCR7– memory T cells do not differ in immediate effector cell function. J. Immunol. 169:638-641.
Wherry, E. J., V. Teichgraber, T. C. Becker, D. Masopust, S. M. Kaech, R. Antia, U. H. von Andrian, and R. Ahmed. 2003. Lineage relationship and protective immunity of memory CD8 T cell subsets. Nat. Immunol. 4:225-234.
Wolint, P., M. R. Betts, R. A. Koup, and A. Oxenius. 2004. Immediate cytotoxicity but not degranulation distinguishes effector and memory subsets of CD8+ T cells. J. Exp. Med. 199:925-936.
Zimmerman, C., K. Brduscha-Riem, C. Blaser, R. M. Zinkernagel, and H. Pircher. 1996. Visualization, characterization, and turnover of CD8+ memory T cells in virus-infected hosts. J. Exp. Med. 183:1367-1375.
Zimmermann, C., A. Prevost-Blondel, C. Blaser, and H. Pircher. 1999. Kinetics of the response of naive and memory CD8 T cells to antigen: similarities and differences. Eur. J. Immunol. 29:284-290.(Heike Unsoeld and Hanspet)