The adenylate cyclase toxin of Bacillus anthracis is a potent promoter of TH17 cell development
2011; Elsevier BV; Volume: 127; Issue: 6 Linguagem: Inglês
10.1016/j.jaci.2010.12.1104
ISSN1097-6825
AutoresSilvia Rossi Paccani, Marisa Benagiano, Maria Teresa Savino, Francesca Finetti, Fiorella Tonello, Mario Milco D’Elios, Cosima T. Baldari,
Tópico(s)SARS-CoV-2 and COVID-19 Research
ResumoTo the Editor:Pathogenic strains of Bacillus anthracis produce 2 toxins: lethal factor, a metalloproteinase that cleaves mitogen-activated protein kinase kinases, and edema factor (EF), a Ca2+/calmodulin-dependent adenylate cyclase, the entry of which into target cells requires their assembly as binary complexes with protective antigen (PA).1Collier R.J. Membrane translocation by anthrax toxin.Mol Aspects Med. 2009; 30: 413-422Crossref PubMed Scopus (119) Google Scholar By targeting crucial components of the signaling pathways implicated in the activation of both innate and adaptive immunity, these toxins cooperatively suppress the immune response and hence provide B anthracis with a powerful means of immune evasion in the critical initial phase of infection.2Tournier J.N. Rossi Paccani S. Quesnel-Hellmann A. Baldari C.T. Anthrax toxins: a weapon to systematically dismantle the host immune defenses.Mol Aspects Med. 2009; 30: 456-466Crossref PubMed Scopus (101) Google Scholar We have recently shown that T cells are exquisitely sensitive to edema toxin (ET; EF+PA), such that at low nonimmunosuppressive concentrations, the toxin selectively targets specific signaling modules in the T-cell receptor (TCR) signaling cascade through its cyclic AMP (cAMP)–increasing activity, thereby promoting TH2 cell development.3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google ScholarOn encounter with antigen exposed on antigen-presenting cells (APCs), naive conventional CD4+ T cells enter a complex differentiation program, which leads to their eventual development into effectors with distinct and complementary activities, with the TH1 and TH17 subsets responsible for orchestrating the cell-mediated immune response to the invading pathogen and the TH2 subset responsible for optimizing the humoral response. A bias toward the TH1 and, to an even larger extent, the TH17 inflammatory subsets has been causally associated with the pathogenesis of a number of autoimmune diseases.4Murphy K.M. Stockinger B. Effector T cell plasticity: flexibility in the face of changing circumstances.Nat Immunol. 2010; 11: 674-680Crossref PubMed Scopus (350) Google Scholar As such, the balance among TH subsets is considered an important target in both vaccination and therapeutic intervention.Naive CD4+ T-cell priming to a specific TH lineage is crucially dependent on environmental cues, which include cytokines and lipid mediators produced by APCs in response to the specific features of the invading pathogen. Among the latter, prostaglandin E2, which has the ability to skew the TH1/TH2 balance toward the TH2 subset, has been recently reported to also favor TH17 cell development through the cAMP-dependent upregulation of IL-23 and IL-1β receptor expression,5Boniface K. Bak-Jensen K.S. Li Y. Blumenschein W.M. McGeachy M.J. McClanahan T.K. et al.Prostaglandin E2 regulates Th17 cell differentiation and function through cyclic AMP and EP2/EP4 receptor signaling.J Exp Med. 2009; 206: 535-548Crossref PubMed Scopus (349) Google Scholar further highlighting the multifaceted role of cAMP as an immunomodulator. Here we have assessed the potential effects of the anthrax adenylate cyclase toxin on TH17 cell development.Enriched CD4+ T cells purified from buffy coats from healthy donors by using the StemSep Human T-cell enrichment kit (Voden Medical Instruments, Milan, Italy) were activated by anti-CD3 mAb (OKT3) after preincubation, where required, with ET (0.11 nmol/L; PA/EF ratio, 1.6) or CyaA (0.28 nmol/L) or the respective adenylate cyclase–defective mutants EL1 and CyaA-E5, as previously described,3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google Scholar for 2 hours. Recombinant human IL-2 (30 U/mL; Eurocetus, Milan, Italy) was added to the cultures on days 4 and 7. After 10 days, cells were washed and restimulated for 24 hours, 36 hours, or both with anti-CD3 mAb, and cytokines were measured by using ELISPOT, ELISA, or both. RNA extraction and real-time quantitative RT-PCR were performed as previously described3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google Scholar by using the SYBR Green I SensiMix dT Kit (Quantace, Watford, United Kingdom).Priming of CD4+ T cells in neutral conditions (anti-CD3 mAb, no cytokines) in the presence of a low ET concentration (0.11 nmol/L) that does not affect proliferation but is sufficient to modulate TCR signaling3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google Scholar results in a robust expansion of TH17 cells, as assessed by IL-17, IL-22, and IL-21 production (Fig 1, A). Consistent with these findings, the amount of mRNA encoding retinoic acid receptor–related orphan receptor γ, the master transcription factor in TH17 differentiation,4Murphy K.M. Stockinger B. Effector T cell plasticity: flexibility in the face of changing circumstances.Nat Immunol. 2010; 11: 674-680Crossref PubMed Scopus (350) Google Scholar was enhanced under these conditions (Fig 1, A). Real-time RT-PCR analysis revealed that ET treatment also resulted in an increased expression of IL-23 and IL-1 receptors (Fig 1, A), which are upregulated in TH17 cells.4Murphy K.M. Stockinger B. Effector T cell plasticity: flexibility in the face of changing circumstances.Nat Immunol. 2010; 11: 674-680Crossref PubMed Scopus (350) Google Scholar The levels of IL-10, which is produced by human TH17 cells (but also by other TH subsets) as a mechanism to limit the magnitude of the immune response and prevent tissue damage,4Murphy K.M. Stockinger B. Effector T cell plasticity: flexibility in the face of changing circumstances.Nat Immunol. 2010; 11: 674-680Crossref PubMed Scopus (350) Google Scholar were only modestly increased under these conditions (not shown). Consistent with our previous report,3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google Scholar ET also favors TH2 cell polarization, as shown by its capacity to enhance IL-4 while impairing IFN-γ production (Fig 1, B and C). No effect of ET on the development of regulatory T cells was observed, as assessed by the percentage of CD4+ cells expressing forkhead box protein 3 (data not shown). The TH17-polarizing effects of ET are mediated by cAMP because they were not reproduced by the adenylate cyclase–deficient mutant EL1 (Fig 1, A). To further substantiate these findings, we used an unrelated bacterial adenylase cyclase toxin, Bordetella pertussis CyaA. Similar to ET, nonimmunosuppressive concentrations of CyaA (0.28 nmol/L)3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google Scholar but not of the adenylate cyclase–deficient CyaA-E5 mutant promoted TH17 (and TH2) cell development (Fig 1). Together these findings provide strong evidence that the TH17-polarizing activity of ET depends on its capacity to increase the levels of intracellular cAMP.cAMP acts as a potent immunosuppressor. In T cells this activity involves impairing not only the first step in TCR signaling (ie, activation of the initiating kinase Lck) but also major downstream signaling mediators, such as the mitogen-activating protein kinase cascade.6Tasken K. Stokka A.J. The molecular machinery for cAMP-dependent immunomodulation in T-cells.Biochem Soc Trans. 2006; 34: 476-479Crossref PubMed Scopus (8) Google Scholar Depending on the local concentrations, cAMP can, however, also affect the TH1/TH2 balance, both indirectly by modulating the pattern of cytokines produced by APCs7Sakata D. Yao C. Narumiya S. Prostaglandin E(2), an immunoactivator.J Pharmacol Sci. 2010; 112: 1-5Crossref PubMed Scopus (97) Google Scholar and directly by shaping TCR signaling,3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google Scholar resulting in the selective polarization of CD4+ T cells to the TH2 subset. Using ET and CyaA to increase intracellular cAMP, we have previously reported that cAMP selectively targets specific nodes in the TCR signaling cascade (Akt1, Vav1, and p38) crucial to TH1/TH2 lineage commitment, which results in expression of TH2 transcription factors and development of CD4+ T cells to TH2 effectors, with a concomitant impairment in TH1 cell development.3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google Scholar Here we show that ET and CyaA also favor TH17 cell development. The data not only identify a novel immunomodulatory activity of this toxin but also provide strong evidence of the instructive role of cAMP on the TH17 lineage. Consistent with this notion, other biological molecules capable of enhancing cAMP production, such as prostaglandin E2, were shown to promote TH17 cell development, both by affecting cytokine production by APCs (eg, IL-1β and IL-23) and by targeting T cells through the EP2/EP4 receptors.5Boniface K. Bak-Jensen K.S. Li Y. Blumenschein W.M. McGeachy M.J. McClanahan T.K. et al.Prostaglandin E2 regulates Th17 cell differentiation and function through cyclic AMP and EP2/EP4 receptor signaling.J Exp Med. 2009; 206: 535-548Crossref PubMed Scopus (349) Google Scholar, 7Sakata D. Yao C. Narumiya S. Prostaglandin E(2), an immunoactivator.J Pharmacol Sci. 2010; 112: 1-5Crossref PubMed Scopus (97) Google ScholarThe mechanisms underlying the mixed TH2/TH17-polarizing activity of cAMP remain to be defined. Under our experimental conditions aimed at assessing the direct effects of cAMP on T cells in the absence of APC-derived cytokines, the most plausible target is the TCR signaling cascade at priming. We could hypothesize a scenario in which cAMP modulates TCR signaling to inhibit the signaling mediators leading to TH1 development while enhancing the activation of distinct signaling mediators implicated in commitment to the TH2 and TH17 lineages, respectively. The activation might be stochastic, resulting in the development of individual cells into one or the other of these 2 subsets. In the physiologic context of antigen presentation, this decision will be reinforced or modified by the APC-derived cytokines. Alternatively, cAMP might favor commitment to only one of the 2 lineages, but developing TH17 cells, which are characterized by a remarkable plasticity, as well as instability,4Murphy K.M. Stockinger B. Effector T cell plasticity: flexibility in the face of changing circumstances.Nat Immunol. 2010; 11: 674-680Crossref PubMed Scopus (350) Google Scholar could switch to the TH2 lineage during their differentiation. It should be pointed out that although TH17 cells produce IL-22, in human subjects a major cellular source of this cytokine is represented by the newly identified TH22 subset.4Murphy K.M. Stockinger B. Effector T cell plasticity: flexibility in the face of changing circumstances.Nat Immunol. 2010; 11: 674-680Crossref PubMed Scopus (350) Google Scholar The observation that ET and CyaA upregulate IL-22 production suggests the interesting possibility that cAMP might have an even wider-ranging instructive role in TH cell development.Although the key role of TH2 cells in the development of antibody-mediated immunity is well established and is in fact exploited to generate effective vaccine adjuvants, the importance of TH17 cells not only for primary immune responses but also for vaccine-induced memory responses against infectious diseases has been only recently appreciated.8Lin Y. Slight S.R. Khader S.A. Th17 cytokines and vaccine-induced immunity.Semin Immunopathol. 2010; 32: 79-90Crossref PubMed Scopus (89) Google Scholar When coadministered with antigen to mice, ET potentiates antibody responses by favoring the development of antigen-specific TH2 cells.9Duverger A. Jackson R.J. van Ginkel F.W. Fischer R. Tafaro A. Leppla S.H. et al.Bacillus anthracis edema toxin acts as an adjuvant for mucosal immune responses to nasally administered vaccine antigens.J Immunol. 2006; 176: 1776-1783PubMed Google Scholar Our finding that ET is also a potent inducer of TH17 cells, an effect that is reproduced by CyaA, suggests that cAMP might enhance the efficacy of vaccines by acting on both the humoral and cellular arms of adaptive immunity. This TH2/TH17-polarizing activity of ET might likely account for the robust humoral and cell-mediated immunity in the bioterrorism-related anthrax exposures occurring at the US Capitol in 2001. To the Editor: Pathogenic strains of Bacillus anthracis produce 2 toxins: lethal factor, a metalloproteinase that cleaves mitogen-activated protein kinase kinases, and edema factor (EF), a Ca2+/calmodulin-dependent adenylate cyclase, the entry of which into target cells requires their assembly as binary complexes with protective antigen (PA).1Collier R.J. Membrane translocation by anthrax toxin.Mol Aspects Med. 2009; 30: 413-422Crossref PubMed Scopus (119) Google Scholar By targeting crucial components of the signaling pathways implicated in the activation of both innate and adaptive immunity, these toxins cooperatively suppress the immune response and hence provide B anthracis with a powerful means of immune evasion in the critical initial phase of infection.2Tournier J.N. Rossi Paccani S. Quesnel-Hellmann A. Baldari C.T. Anthrax toxins: a weapon to systematically dismantle the host immune defenses.Mol Aspects Med. 2009; 30: 456-466Crossref PubMed Scopus (101) Google Scholar We have recently shown that T cells are exquisitely sensitive to edema toxin (ET; EF+PA), such that at low nonimmunosuppressive concentrations, the toxin selectively targets specific signaling modules in the T-cell receptor (TCR) signaling cascade through its cyclic AMP (cAMP)–increasing activity, thereby promoting TH2 cell development.3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google Scholar On encounter with antigen exposed on antigen-presenting cells (APCs), naive conventional CD4+ T cells enter a complex differentiation program, which leads to their eventual development into effectors with distinct and complementary activities, with the TH1 and TH17 subsets responsible for orchestrating the cell-mediated immune response to the invading pathogen and the TH2 subset responsible for optimizing the humoral response. A bias toward the TH1 and, to an even larger extent, the TH17 inflammatory subsets has been causally associated with the pathogenesis of a number of autoimmune diseases.4Murphy K.M. Stockinger B. Effector T cell plasticity: flexibility in the face of changing circumstances.Nat Immunol. 2010; 11: 674-680Crossref PubMed Scopus (350) Google Scholar As such, the balance among TH subsets is considered an important target in both vaccination and therapeutic intervention. Naive CD4+ T-cell priming to a specific TH lineage is crucially dependent on environmental cues, which include cytokines and lipid mediators produced by APCs in response to the specific features of the invading pathogen. Among the latter, prostaglandin E2, which has the ability to skew the TH1/TH2 balance toward the TH2 subset, has been recently reported to also favor TH17 cell development through the cAMP-dependent upregulation of IL-23 and IL-1β receptor expression,5Boniface K. Bak-Jensen K.S. Li Y. Blumenschein W.M. McGeachy M.J. McClanahan T.K. et al.Prostaglandin E2 regulates Th17 cell differentiation and function through cyclic AMP and EP2/EP4 receptor signaling.J Exp Med. 2009; 206: 535-548Crossref PubMed Scopus (349) Google Scholar further highlighting the multifaceted role of cAMP as an immunomodulator. Here we have assessed the potential effects of the anthrax adenylate cyclase toxin on TH17 cell development. Enriched CD4+ T cells purified from buffy coats from healthy donors by using the StemSep Human T-cell enrichment kit (Voden Medical Instruments, Milan, Italy) were activated by anti-CD3 mAb (OKT3) after preincubation, where required, with ET (0.11 nmol/L; PA/EF ratio, 1.6) or CyaA (0.28 nmol/L) or the respective adenylate cyclase–defective mutants EL1 and CyaA-E5, as previously described,3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google Scholar for 2 hours. Recombinant human IL-2 (30 U/mL; Eurocetus, Milan, Italy) was added to the cultures on days 4 and 7. After 10 days, cells were washed and restimulated for 24 hours, 36 hours, or both with anti-CD3 mAb, and cytokines were measured by using ELISPOT, ELISA, or both. RNA extraction and real-time quantitative RT-PCR were performed as previously described3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google Scholar by using the SYBR Green I SensiMix dT Kit (Quantace, Watford, United Kingdom). Priming of CD4+ T cells in neutral conditions (anti-CD3 mAb, no cytokines) in the presence of a low ET concentration (0.11 nmol/L) that does not affect proliferation but is sufficient to modulate TCR signaling3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google Scholar results in a robust expansion of TH17 cells, as assessed by IL-17, IL-22, and IL-21 production (Fig 1, A). Consistent with these findings, the amount of mRNA encoding retinoic acid receptor–related orphan receptor γ, the master transcription factor in TH17 differentiation,4Murphy K.M. Stockinger B. Effector T cell plasticity: flexibility in the face of changing circumstances.Nat Immunol. 2010; 11: 674-680Crossref PubMed Scopus (350) Google Scholar was enhanced under these conditions (Fig 1, A). Real-time RT-PCR analysis revealed that ET treatment also resulted in an increased expression of IL-23 and IL-1 receptors (Fig 1, A), which are upregulated in TH17 cells.4Murphy K.M. Stockinger B. Effector T cell plasticity: flexibility in the face of changing circumstances.Nat Immunol. 2010; 11: 674-680Crossref PubMed Scopus (350) Google Scholar The levels of IL-10, which is produced by human TH17 cells (but also by other TH subsets) as a mechanism to limit the magnitude of the immune response and prevent tissue damage,4Murphy K.M. Stockinger B. Effector T cell plasticity: flexibility in the face of changing circumstances.Nat Immunol. 2010; 11: 674-680Crossref PubMed Scopus (350) Google Scholar were only modestly increased under these conditions (not shown). Consistent with our previous report,3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google Scholar ET also favors TH2 cell polarization, as shown by its capacity to enhance IL-4 while impairing IFN-γ production (Fig 1, B and C). No effect of ET on the development of regulatory T cells was observed, as assessed by the percentage of CD4+ cells expressing forkhead box protein 3 (data not shown). The TH17-polarizing effects of ET are mediated by cAMP because they were not reproduced by the adenylate cyclase–deficient mutant EL1 (Fig 1, A). To further substantiate these findings, we used an unrelated bacterial adenylase cyclase toxin, Bordetella pertussis CyaA. Similar to ET, nonimmunosuppressive concentrations of CyaA (0.28 nmol/L)3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google Scholar but not of the adenylate cyclase–deficient CyaA-E5 mutant promoted TH17 (and TH2) cell development (Fig 1). Together these findings provide strong evidence that the TH17-polarizing activity of ET depends on its capacity to increase the levels of intracellular cAMP. cAMP acts as a potent immunosuppressor. In T cells this activity involves impairing not only the first step in TCR signaling (ie, activation of the initiating kinase Lck) but also major downstream signaling mediators, such as the mitogen-activating protein kinase cascade.6Tasken K. Stokka A.J. The molecular machinery for cAMP-dependent immunomodulation in T-cells.Biochem Soc Trans. 2006; 34: 476-479Crossref PubMed Scopus (8) Google Scholar Depending on the local concentrations, cAMP can, however, also affect the TH1/TH2 balance, both indirectly by modulating the pattern of cytokines produced by APCs7Sakata D. Yao C. Narumiya S. Prostaglandin E(2), an immunoactivator.J Pharmacol Sci. 2010; 112: 1-5Crossref PubMed Scopus (97) Google Scholar and directly by shaping TCR signaling,3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google Scholar resulting in the selective polarization of CD4+ T cells to the TH2 subset. Using ET and CyaA to increase intracellular cAMP, we have previously reported that cAMP selectively targets specific nodes in the TCR signaling cascade (Akt1, Vav1, and p38) crucial to TH1/TH2 lineage commitment, which results in expression of TH2 transcription factors and development of CD4+ T cells to TH2 effectors, with a concomitant impairment in TH1 cell development.3Rossi Paccani S. Benagiano M. Capitani N. Zornetta I. Ladant D. Montecucco C. et al.The adenylate cyclase toxins of Bacillus anthracis and Bordetella pertussis promote Th2 cell development by shaping T cell antigen receptor signaling.PLoS Pathog. 2009; 5: e1000325Crossref PubMed Scopus (42) Google Scholar Here we show that ET and CyaA also favor TH17 cell development. The data not only identify a novel immunomodulatory activity of this toxin but also provide strong evidence of the instructive role of cAMP on the TH17 lineage. Consistent with this notion, other biological molecules capable of enhancing cAMP production, such as prostaglandin E2, were shown to promote TH17 cell development, both by affecting cytokine production by APCs (eg, IL-1β and IL-23) and by targeting T cells through the EP2/EP4 receptors.5Boniface K. Bak-Jensen K.S. Li Y. Blumenschein W.M. McGeachy M.J. McClanahan T.K. et al.Prostaglandin E2 regulates Th17 cell differentiation and function through cyclic AMP and EP2/EP4 receptor signaling.J Exp Med. 2009; 206: 535-548Crossref PubMed Scopus (349) Google Scholar, 7Sakata D. Yao C. Narumiya S. Prostaglandin E(2), an immunoactivator.J Pharmacol Sci. 2010; 112: 1-5Crossref PubMed Scopus (97) Google Scholar The mechanisms underlying the mixed TH2/TH17-polarizing activity of cAMP remain to be defined. Under our experimental conditions aimed at assessing the direct effects of cAMP on T cells in the absence of APC-derived cytokines, the most plausible target is the TCR signaling cascade at priming. We could hypothesize a scenario in which cAMP modulates TCR signaling to inhibit the signaling mediators leading to TH1 development while enhancing the activation of distinct signaling mediators implicated in commitment to the TH2 and TH17 lineages, respectively. The activation might be stochastic, resulting in the development of individual cells into one or the other of these 2 subsets. In the physiologic context of antigen presentation, this decision will be reinforced or modified by the APC-derived cytokines. Alternatively, cAMP might favor commitment to only one of the 2 lineages, but developing TH17 cells, which are characterized by a remarkable plasticity, as well as instability,4Murphy K.M. Stockinger B. Effector T cell plasticity: flexibility in the face of changing circumstances.Nat Immunol. 2010; 11: 674-680Crossref PubMed Scopus (350) Google Scholar could switch to the TH2 lineage during their differentiation. It should be pointed out that although TH17 cells produce IL-22, in human subjects a major cellular source of this cytokine is represented by the newly identified TH22 subset.4Murphy K.M. Stockinger B. Effector T cell plasticity: flexibility in the face of changing circumstances.Nat Immunol. 2010; 11: 674-680Crossref PubMed Scopus (350) Google Scholar The observation that ET and CyaA upregulate IL-22 production suggests the interesting possibility that cAMP might have an even wider-ranging instructive role in TH cell development. Although the key role of TH2 cells in the development of antibody-mediated immunity is well established and is in fact exploited to generate effective vaccine adjuvants, the importance of TH17 cells not only for primary immune responses but also for vaccine-induced memory responses against infectious diseases has been only recently appreciated.8Lin Y. Slight S.R. Khader S.A. Th17 cytokines and vaccine-induced immunity.Semin Immunopathol. 2010; 32: 79-90Crossref PubMed Scopus (89) Google Scholar When coadministered with antigen to mice, ET potentiates antibody responses by favoring the development of antigen-specific TH2 cells.9Duverger A. Jackson R.J. van Ginkel F.W. Fischer R. Tafaro A. Leppla S.H. et al.Bacillus anthracis edema toxin acts as an adjuvant for mucosal immune responses to nasally administered vaccine antigens.J Immunol. 2006; 176: 1776-1783PubMed Google Scholar Our finding that ET is also a potent inducer of TH17 cells, an effect that is reproduced by CyaA, suggests that cAMP might enhance the efficacy of vaccines by acting on both the humoral and cellular arms of adaptive immunity. This TH2/TH17-polarizing activity of ET might likely account for the robust humoral and cell-mediated immunity in the bioterrorism-related anthrax exposures occurring at the US Capitol in 2001. We thank Dr Daniel Ladant for his kind gift of CyaA and CyaA-E5, as well as for his helpful comments, and Drs Cesare Montecucco and John L. Telford for useful discussions and critical reading of the manuscript.
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