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Conditioning of Dendritic Cells by Pathogen-Derived Stimuli
Immunobiol. (2001) 204, pp. 595 – 597 © 2001 Urban & Fischer Verlag http://www.urbanfischer.de/journals/immunobiol
Immunobiology Laboratory, Imperial Cancer Research Fund, London, UK
Conditioning of Dendritic Cells by Pathogen-Derived Stimuli CAETANO REIS E SOUSA, ALEXANDER D. EDWARDS, SHIVANTHI P. MANICKASINGHAM, and OLIVER SCHULZ
Dendritic cells (DC) are among the most powerful regulators of adaptive immunity. They are present in most tissues in a quiescent or resting state in which they have limited ability to interact with T cells until they become “activated” and transit to an “effector” state. There appear to be multiple types of effector DC, ranging from tolerogenic DC to immunogenic DC favouring Th1 (“DC1”) or Th2 responses (“DC2”). The differences in the function of these cell types may be due to quantitative or qualitative differences in DC activation or to functional specialisation of certain DC subtypes (1–4). It is widely recognized that infection is a major stimulus for generating immunogenic DC. Indeed, DC appear to be specialized to recognize and become activated by a wide variety of hallmarks of infection. These include microbial structures, proteins released by damaged cells and inflammatory mediators produced by infected or bystander cells (5). As a paradigm for understanding DC activation in a physiological context, we have been studying the regulation of cytokine production by DC in response to microbial recognition. Using a combination of in vivo and in vitro models, we have found that murine splenic DC exposed to extracts of Toxoplasma gondii (STAg), CpG-containing DNA oligonucleotides, heat-killed Listeria monocytogenes or Brucella abortus rapidly produce the Th1-promoting cytokine, IL-12 (6-9). However, production of IL-12 elicited by these agents is biased towards the IL-12 p40 subunit and, for many of the stimuli, production of bioactive IL-12 p70 heterodimer is low or undetectable. However, these stimuli all elicit high levels of IL-12 p70 from DC when combined with CD40 cross-linking, both in vitro and in vivo (7) (Fig. 1). CD40 triggering mimics signals delivered to DC by T cells during cognate DC: T cell interactions and IL-12 p70 is also produced by DC in an in vitro model of naïve transgenic T cell activation when these microbial products are added as co-stimuli (7, 9). Importantly, neither CD40 crosslinking in vivo nor T cell feedback in vitro can initiate IL-12 production. Injection of mice with LPS-free anti-CD40 mAbs fails to trigger production of either IL-12 p40 or IL-12 p35 by splenic DC (7). Thus, the CD40/T cell feedback pathway only acts to amplify IL-12 production by DC conditioned by a microbial stimulus. T cell feedback and CD40 cross-linking appear to potentiate IL-12 p70 production by increasing expression of IL-12 p35 (7). Not all microbial activators of DC have this effect. Indeed, we have tested a large number of putative DC activators and found that many are able to trigger DC activation as measured by an upregulation of co-stimulatory ability but do not condition the cells to produce IL-12 (Edwards et al, unpublished). However, despite the fact that they do not 0171-2985/01/204/05-595 $ 15.00/0
596 · C. REIS E SOUSA et al.
Figure 1.
trigger production of DC cytokines by themselves, some stimuli such as heat-killed yeast or zymosan particles condition DC to produce IL-10 in response to CD40 triggering or T cell feedback (9) (Fig. 1). This suggests that the CD40 pathway in DC is not “wired” to elicit IL-12 production. As might be expected, DC conditioned to make IL-12 promote Th1 development in an in vitro model of differentiation using naïve transgenic T cells. In contrast, DC conditioned by yeasts to make IL-10 suppress Th1 development and allow emergence of Th2 cells. These effects can be abrogated by neutralisation of IL-12 or IL-10, respectively, establishing the link between cytokine production by DC and subsequent T cell differentiation (9). Thus, the nature of the microbial activator dictates the type of immunomodulatory cytokine produced in response to CD40 signalling. All of these experiments make use of heterogeneous populations of DC. Mouse spleen DC contain at least 3 distinct sub-populations defined on the basis of CD4 and CD8a expression. CD4+, CD4– CD8a– (DN) and CD8a+ DC all appear to constitute stable DC subsets with differing cytokine production potential, and, possibly, distinct functions (10). The question of whether DC subsets are pre-committed “DC1“ and “DC2“ or respond in a flexible manner to environmental cues is a matter of debate (3–5). It has been argued that CD8a+ DC are unique in their ability to make IL-12 and promote Th1 responses while other subsets promote Th2-dominated immunity (11, 12). In contrast, the ability of DC subsets to make IL-10 has not been studied in detail. These questions led us to examine the cytokine production patterns of purified DC subsets in response to IL-10- or IL-12- promoting stimuli. None of the subsets make high levels of IL-10- or IL-12-p70 spontaneously but all can be triggered by microbial stimulation plus CD40 ligation. Interestingly, CD8a+ and DN DC can make either IL-12 p70 or IL-10 in response to the appropriate stimulus. These results argue in favour of the hypothesis that many DC types are flexible and adjust their effector function in response to the type of pathogen encountered, although it remains possible that unaccounted for heterogeneity
Regulation of DC by pathogens · 597
in CD8a+ and DN DC populations is responsible for differential IL-10 and IL-12 production. In contrast to these results with CD8a+ and DN DC, CD4+ DC appear to be unable to make IL-12 p70 in response to any stimulus. Thus, some DC subsets may be display functional specialisation. In summary, our results demonstrate that the function of immunogenic DC is largely dictated by microbial encounter. DC are resting until they receive signals that signify the presence of infection. During subsequent interactions, T cells deliver signals that potently amplify DC function but do not alter the quality of DC activation. Because signals from T cells fail to initiate de novo cytokine production by DC, the latter remains firmly under control of the innate immune system. These properties of DC allow this specialised group of APC to establish a unique link between innate recognition and adaptive immunity.
References 1. BANCHEREAU, J., F. BRIERE, C. CAUX, J. DAVOUST, S. LEBECQUE, Y. J. LIU, B. PULENDRAN, and K. PALUCKA. 2000. Immunobiology of dendritic cells. Annu. Rev. Immunol. 18: 767–811. 2. MOSER, M., and K. M. MURPHY. 2000. Dendritic cell regulation of THI–TH2 development. Nature Immunol. 1: 199 205. 3. LANZAVECCHIA, A., and F. SALLUSTO. 2001. The instructive role of dendritic cells on T cell responses: lineages, plasticity and kinetics. Curr. Opin. Immunol. 13: 291–298. 4. LIU, Y. J., H. KANZLER, V. SOUMELIS, and M. GILLIET. 2001. Dendritic cell lineage, plasticity and cross-regulation. Nat. Immunol. 2: 585–589. 5. REIS E SOUSA, C. 2001. Dendritic cells as sensors of infection. Immunity 14: 495–498. 6. REIS E SOUSA, C., S. HIENY, T. SCHARTON-KERSTEN, H. CHAREST, D. JANKOVIC, R. N. GERMAIN, and A. SHER. 1997. In vivo microbial stimulation induces rapid CD40L-independent production of IL-12 by dendritic cells and their re-distribution to T cell areas. J. Exp. Med. 186: 1819–1829. 7. SCHULZ, O., A. D. EDWARDS, M. SCHITO, J. ALIBERTI, S. MANICKASINGHAM, A. SHER, and C. REIS E SOUSA. 2000. CD40 triggering of heterodimeric IL-12 p70 production by dendritic cells in vivo requires a microbial priming signal. Immunity 13: 453–462. 8. HUANG, L.-Y., C. REIS E SOUSA, Y. ITOH, J. INMAN, and D. E. SCOTT. 2001. IL-12 induction by a Th1-Inducing adjuvant in vivo: dendritic cell subsets and regulation by IL-10. J. Immunol. 167: 1423–1430. 9. EDWARDS, A. D., S. P. MANICKASINGHAM, R. SPÖRRI, S. S. DIEBOLD, O. SCHULZ, A. SHER, T. KAISHO, S. AKIRA, and C. REIS E SOUSA. 2001. Microbial recognition via toll-like receptor-dependent and -independent pathways determines the cytokine response of murine dendritic cell subsets to CD40 triggering. Submitted. 10. SHORTMAN, K. 2000. Burnet oration: dendritic cells: multiple subtypes, multiple origins, multiple functions. Immunol. Cell. Biol. 78: 161–165. 11. PULENDRAN, B., J. L. SMITH, G. CASPARY, K. BRASEL, D. PETTIT, E. MARASKOVSKY, and C. R. MALISZEWSKI. 1999. Distinct dendritic cell subsets differentially regulate the class of immune response in vivo. Proc. Natl. Acad. Sci. USA 96: 1036–1041. 12. MALDONADO-LÓPEZ, R., T. DE SMEDT, P. MICHEL, J. GODFROID, B. PAJAK, C. HEIRMAN, K. THIELEMANS, O. LEO, J. URBAIN, and M. MOSER. 1999. CD8a+ and CD8a– Subclasses of Dendritic Cells Direct the Development of Distinct T Helper Cells In Vivo. J. Exp. Med. 189: 587–592. Dr. CAETANO REIS E SOUSA, Immunobiology Laboratory, Imperial Cancer Research Fund, 44 Lincoln’s Inn Fields, London WC2A 3PX, United Kingdom, Tel.: +44 20 7269 2832, Fax: +44 20 7269 2833, E-mail: [email protected]
Immunobiology Laboratory, Imperial Cancer Research Fund, London, UK
Conditioning of Dendritic Cells by Pathogen-Derived Stimuli CAETANO REIS E SOUSA, ALEXANDER D. EDWARDS, SHIVANTHI P. MANICKASINGHAM, and OLIVER SCHULZ
Dendritic cells (DC) are among the most powerful regulators of adaptive immunity. They are present in most tissues in a quiescent or resting state in which they have limited ability to interact with T cells until they become “activated” and transit to an “effector” state. There appear to be multiple types of effector DC, ranging from tolerogenic DC to immunogenic DC favouring Th1 (“DC1”) or Th2 responses (“DC2”). The differences in the function of these cell types may be due to quantitative or qualitative differences in DC activation or to functional specialisation of certain DC subtypes (1–4). It is widely recognized that infection is a major stimulus for generating immunogenic DC. Indeed, DC appear to be specialized to recognize and become activated by a wide variety of hallmarks of infection. These include microbial structures, proteins released by damaged cells and inflammatory mediators produced by infected or bystander cells (5). As a paradigm for understanding DC activation in a physiological context, we have been studying the regulation of cytokine production by DC in response to microbial recognition. Using a combination of in vivo and in vitro models, we have found that murine splenic DC exposed to extracts of Toxoplasma gondii (STAg), CpG-containing DNA oligonucleotides, heat-killed Listeria monocytogenes or Brucella abortus rapidly produce the Th1-promoting cytokine, IL-12 (6-9). However, production of IL-12 elicited by these agents is biased towards the IL-12 p40 subunit and, for many of the stimuli, production of bioactive IL-12 p70 heterodimer is low or undetectable. However, these stimuli all elicit high levels of IL-12 p70 from DC when combined with CD40 cross-linking, both in vitro and in vivo (7) (Fig. 1). CD40 triggering mimics signals delivered to DC by T cells during cognate DC: T cell interactions and IL-12 p70 is also produced by DC in an in vitro model of naïve transgenic T cell activation when these microbial products are added as co-stimuli (7, 9). Importantly, neither CD40 crosslinking in vivo nor T cell feedback in vitro can initiate IL-12 production. Injection of mice with LPS-free anti-CD40 mAbs fails to trigger production of either IL-12 p40 or IL-12 p35 by splenic DC (7). Thus, the CD40/T cell feedback pathway only acts to amplify IL-12 production by DC conditioned by a microbial stimulus. T cell feedback and CD40 cross-linking appear to potentiate IL-12 p70 production by increasing expression of IL-12 p35 (7). Not all microbial activators of DC have this effect. Indeed, we have tested a large number of putative DC activators and found that many are able to trigger DC activation as measured by an upregulation of co-stimulatory ability but do not condition the cells to produce IL-12 (Edwards et al, unpublished). However, despite the fact that they do not 0171-2985/01/204/05-595 $ 15.00/0
596 · C. REIS E SOUSA et al.
Figure 1.
trigger production of DC cytokines by themselves, some stimuli such as heat-killed yeast or zymosan particles condition DC to produce IL-10 in response to CD40 triggering or T cell feedback (9) (Fig. 1). This suggests that the CD40 pathway in DC is not “wired” to elicit IL-12 production. As might be expected, DC conditioned to make IL-12 promote Th1 development in an in vitro model of differentiation using naïve transgenic T cells. In contrast, DC conditioned by yeasts to make IL-10 suppress Th1 development and allow emergence of Th2 cells. These effects can be abrogated by neutralisation of IL-12 or IL-10, respectively, establishing the link between cytokine production by DC and subsequent T cell differentiation (9). Thus, the nature of the microbial activator dictates the type of immunomodulatory cytokine produced in response to CD40 signalling. All of these experiments make use of heterogeneous populations of DC. Mouse spleen DC contain at least 3 distinct sub-populations defined on the basis of CD4 and CD8a expression. CD4+, CD4– CD8a– (DN) and CD8a+ DC all appear to constitute stable DC subsets with differing cytokine production potential, and, possibly, distinct functions (10). The question of whether DC subsets are pre-committed “DC1“ and “DC2“ or respond in a flexible manner to environmental cues is a matter of debate (3–5). It has been argued that CD8a+ DC are unique in their ability to make IL-12 and promote Th1 responses while other subsets promote Th2-dominated immunity (11, 12). In contrast, the ability of DC subsets to make IL-10 has not been studied in detail. These questions led us to examine the cytokine production patterns of purified DC subsets in response to IL-10- or IL-12- promoting stimuli. None of the subsets make high levels of IL-10- or IL-12-p70 spontaneously but all can be triggered by microbial stimulation plus CD40 ligation. Interestingly, CD8a+ and DN DC can make either IL-12 p70 or IL-10 in response to the appropriate stimulus. These results argue in favour of the hypothesis that many DC types are flexible and adjust their effector function in response to the type of pathogen encountered, although it remains possible that unaccounted for heterogeneity
Regulation of DC by pathogens · 597
in CD8a+ and DN DC populations is responsible for differential IL-10 and IL-12 production. In contrast to these results with CD8a+ and DN DC, CD4+ DC appear to be unable to make IL-12 p70 in response to any stimulus. Thus, some DC subsets may be display functional specialisation. In summary, our results demonstrate that the function of immunogenic DC is largely dictated by microbial encounter. DC are resting until they receive signals that signify the presence of infection. During subsequent interactions, T cells deliver signals that potently amplify DC function but do not alter the quality of DC activation. Because signals from T cells fail to initiate de novo cytokine production by DC, the latter remains firmly under control of the innate immune system. These properties of DC allow this specialised group of APC to establish a unique link between innate recognition and adaptive immunity.
References 1. BANCHEREAU, J., F. BRIERE, C. CAUX, J. DAVOUST, S. LEBECQUE, Y. J. LIU, B. PULENDRAN, and K. PALUCKA. 2000. Immunobiology of dendritic cells. Annu. Rev. Immunol. 18: 767–811. 2. MOSER, M., and K. M. MURPHY. 2000. Dendritic cell regulation of THI–TH2 development. Nature Immunol. 1: 199 205. 3. LANZAVECCHIA, A., and F. SALLUSTO. 2001. The instructive role of dendritic cells on T cell responses: lineages, plasticity and kinetics. Curr. Opin. Immunol. 13: 291–298. 4. LIU, Y. J., H. KANZLER, V. SOUMELIS, and M. GILLIET. 2001. Dendritic cell lineage, plasticity and cross-regulation. Nat. Immunol. 2: 585–589. 5. REIS E SOUSA, C. 2001. Dendritic cells as sensors of infection. Immunity 14: 495–498. 6. REIS E SOUSA, C., S. HIENY, T. SCHARTON-KERSTEN, H. CHAREST, D. JANKOVIC, R. N. GERMAIN, and A. SHER. 1997. In vivo microbial stimulation induces rapid CD40L-independent production of IL-12 by dendritic cells and their re-distribution to T cell areas. J. Exp. Med. 186: 1819–1829. 7. SCHULZ, O., A. D. EDWARDS, M. SCHITO, J. ALIBERTI, S. MANICKASINGHAM, A. SHER, and C. REIS E SOUSA. 2000. CD40 triggering of heterodimeric IL-12 p70 production by dendritic cells in vivo requires a microbial priming signal. Immunity 13: 453–462. 8. HUANG, L.-Y., C. REIS E SOUSA, Y. ITOH, J. INMAN, and D. E. SCOTT. 2001. IL-12 induction by a Th1-Inducing adjuvant in vivo: dendritic cell subsets and regulation by IL-10. J. Immunol. 167: 1423–1430. 9. EDWARDS, A. D., S. P. MANICKASINGHAM, R. SPÖRRI, S. S. DIEBOLD, O. SCHULZ, A. SHER, T. KAISHO, S. AKIRA, and C. REIS E SOUSA. 2001. Microbial recognition via toll-like receptor-dependent and -independent pathways determines the cytokine response of murine dendritic cell subsets to CD40 triggering. Submitted. 10. SHORTMAN, K. 2000. Burnet oration: dendritic cells: multiple subtypes, multiple origins, multiple functions. Immunol. Cell. Biol. 78: 161–165. 11. PULENDRAN, B., J. L. SMITH, G. CASPARY, K. BRASEL, D. PETTIT, E. MARASKOVSKY, and C. R. MALISZEWSKI. 1999. Distinct dendritic cell subsets differentially regulate the class of immune response in vivo. Proc. Natl. Acad. Sci. USA 96: 1036–1041. 12. MALDONADO-LÓPEZ, R., T. DE SMEDT, P. MICHEL, J. GODFROID, B. PAJAK, C. HEIRMAN, K. THIELEMANS, O. LEO, J. URBAIN, and M. MOSER. 1999. CD8a+ and CD8a– Subclasses of Dendritic Cells Direct the Development of Distinct T Helper Cells In Vivo. J. Exp. Med. 189: 587–592. Dr. CAETANO REIS E SOUSA, Immunobiology Laboratory, Imperial Cancer Research Fund, 44 Lincoln’s Inn Fields, London WC2A 3PX, United Kingdom, Tel.: +44 20 7269 2832, Fax: +44 20 7269 2833, E-mail: [email protected]