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The role of CD4 T cell help for CD8 CTL activation
Biochemical and Biophysical Research Communications 384 (2009) 405–408
Contents lists available at ScienceDirect
Biochemical and Biophysical Research Communications journal homepage: www.elsevier.com/locate/ybbrc
Mini Review
The role of CD4 T cell help for CD8 CTL activation Sheng Zhang a,d,*,1, Hongjun Zhang b,1, Jiandong Zhao c a
Medical Oncology, First People’s Hospital, Shanghai Jiaotong University, 650 Xinsongjiang Road, Shanghai 201620, China Medical Oncology, The Medical School Hospital of Qingdao University, Qingdao, China Department of Radiation Oncology, Cancer Hospital of Fudan University, Shanghai, China d Shanghai Key Laboratory of Pancreatic Diseases, Shanghai, China b c
a r t i c l e
i n f o
Article history: Received 19 April 2009 Available online 3 May 2009
a b s t r a c t CD4 T cells play an important role in the initiation and persistence of CD8 T cells responses. In this review, we report on and evaluate the mechanisms by which CD4 T cells contribute to activation of CD8 T cells and the signal pathways of the down-streaming events after CD4 T cell help. Ó 2009 Elsevier Inc. All rights reserved.
Keywords: CD4 T cells CD8 T cells Signal pathway
CD8 CTL cells are essential for protection against viruses, intracellular bacteria infection and tumor cells. Understanding the mechanism for CTL activation, survival and long term maintenance has become one focus for many immunologists. Numerous studies in recent years have reached a consensus that CD4 T cell help is essential for CD8 CTL priming. The help provided by CD4 T lymphocytes during the priming of CD8 T lymphocytes confers a key feature of immune memory: the capacity for autonomous secondary expansion following re-encounter with antigen. Once primed in the presence of CD4 T cells, helped CD8 T cells acquire the ability to undergo a second round of clonal expansion upon restimulation in the absence of T cell help. The CD8 T cells that are not helped by CD4 T cells, in contrast, can mediate effector functions such a s cytotoxicity and cytokine secretion upon restimulation, but do not undergo a second round of clonal expansion [1,2]. These disparate responses have features of being programmed, that is, guided by signals that are transmitted to naïve CD8 T cells during priming, which encode specific fates for their clonal progeny. In this regard, exploring the instructional programme that governs the secondary response of CD8 T cells has significant impact on the future design of CD8 CTL based vaccination. This paper will discuss the mechanisms of CD4 T cell help, the DC licensing model and the downstreamevents following CD4 T cell help pathway.
* Corresponding author. Fax: +86 533 3177425. E-mail address: [email protected] (S. Zhang). 1 These authors contributed equally to this study. 0006-291X/$ - see front matter Ó 2009 Elsevier Inc. All rights reserved. doi:10.1016/j.bbrc.2009.04.134
The mechanisms of CD4 T cell help CD40–CD40L signaling pathway Recent findings have established a DC licensing model. In this model, the CD40L expressing CD4 T cells can interact with the professional APC-DC through the CD40–CD40L pathway. This interaction can lead to the activation/licensing of DC, then the licensed DC can activate CD8 CTL cells. This model was originally proposed/ established by 3 papers in Nature in 1998 [3–5]. In those studies, they found that CTL response was dependent on both CD40 and CD40 ligand and that in the absence of CD4 T helper cells an agonistic antibody to CD40 can substitute for CD4 T cell help. Since the APC was the only known cell to express CD40 and CD4 T helper cells were known to express CD40 ligand upon activation, it was concluded that CD40-dependent licensing of APC was the crucial nature of CD4 T cell help. This proposed licensing model was seriously challenged by another study recently in Science [6]. In this study, the authors could show that CD4 T cell help for the differentiation of CD8 T cells into memory T cells can occur even when all APCs are CD40-deficient. They showed that CD8 T cells can express CD40 transiently after activation. Moreover, CD40-deficient CD8 T cells can never differentiate into memory cells, even in the presence of CD4 T helper cells. They cannot receive helper signals from antigen specific CD4 T cells. Thus, as B cells, the helper signals required for differentiation of CD8 T cells into memory cells appear to pass through the CD40 molecule expressed by activated CD8 T cells. In this model, although CD4 T cell help is CD40-dependent, it is through a direct contact between CD4 T cells and CD8 T cells. The role of DC in this case is to provide the chance that CD4 T cells
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S. Zhang et al. / Biochemical and Biophysical Research Communications 384 (2009) 405–408
and CD8 T cells can be put into proximity for their contact. Thus this study gave direct challenge to the proposed licensing model. Lee and colleagues demonstrated that optimal CD8 CTL response to influenza is dependent on CD40 signaling. However, both primary and secondary response to influenza require CD40 expression on non-T cells [7]. Thus in this case, CD4 T helper cells do not activate CD8 T cells directly through CD40 signaling. With this study, they could show that the direct interaction between CD4 T cells and CD8 T cells through CD40–CD40L signaling is not a universal mechanism for CD4 T cell help. Schuurhuis and colleagues further explored this proposed model. They showed that the DC cell line D1 could activate CD8 CTL with the help of CD4 Th cell [8]. However, when the D1 cells were stimulated with agonistic antiCD40 Ab, they can activate CD8 CTL in a CD4 T cell-independent way, thus further suggesting this DC license model might be true. In 2004, a Nature Immunol paper further gave direct in vivo evidence for DC licensing by Th cells [9]. In this study, they could show that the in vivo sorted DC, only when they were licensed by CD4 T cells, could mediate a memory T cell response. Thus, this model of licensing of DC by CD4 Th cells for CTL priming was finally firmly established.
mediated signal was not identified. It has been reported that signaling of osteoprotgerin ligand (OPGL), another TNF-receptor family member expressed by DC has some similar effects to CD40 signaling. In other words, CD4 T cells can interact with DC through the TRANCE (tumor necrosis factor-related-activation-induced cytokine) pathway which finally leads to the activation of DC [18,19]. Also the CD4 T cells can acquire the effector function for virus protection in this case. Thus TRANCE has been documented as a signaling pathway leading to CD4 T cell activation independent of CD40L [20]. But the use of this signaling pathway by helper cells involved in CTL induction has not been reported. It’s tempting in the future to explore whether this pathway is also used by CD4 T cells for CD8 CTL priming independently of CD40 pathway. The downstreamevents of CD4 T cell help: what happens to DC and CD8 T cells following CD40 ligation? So far, the most characterized and accepted model is still the CD4 Th cells, DC, CD8 CTL model and CD40–CD40L pathway is the major signaling pathway between CD4 Th cells and DC. One important problem would be to address what happened to DC after CD4 T cell licensing so that DC can activate CTL?
IL-2 IL-12 One important characteristic of CD4 T cells is that they can produce multiple cytokines. Thus cytokines have lone been suggested that they can be the mediators of CD4 T cell help. In vitro studies have found that IL-2 is essential for the effective generation of CTL response when helper T cells and CTL were combined in the culture flask [10]. However, conflicting results exist since CTL response to viruses can be generated in IL-2 knock out mice [11]. More recently, using a graft versus host disease model, it was found that the ability of CD4 T helper cells to produce IL-2 was essential for sustained expansion of alloreactive CTL [12]. Using a cross-priming model, several recent studies have demonstrated that the absence of CD4 T cell help for CD8 CTL priming can be overcome by the provision of exogenous IL-2, thus establishing the role of IL-2 for CD4 T cell help [13,14]). More studies need to be done in order to confirm the role of IL-2 as the helper of CD8 CTL or this role can only exist in some certain cases. IFN-c IFN-c is one of the most important type 1 cytokines. It has been shown that IFN-c can enhance the cytotoxicity of CTL in addition to being one of the important effector arms for CD8 CTL. In a recent report, Wolchok and colleagues demonstrated that in a DNA vaccination model, the CTL is fully dependent on CD4 T cell help. However, this CD4 T cell-dependent CTL is lost in IFN-c / mice. It can be restored when recombinant IFN-c was injected in vivo at the priming stage [15], suggesting IFN-c is the essential mediator for CD4 T cell help in this model. A more recent study by Kumaraguru et al. used the well characterized HSV and OVA model system. They found that the helper function of CD4 T cells is mainly mediated by IFN-c producing CD4 Th cells but not IL-2 producing CD4 Th cells [16]. Thus the role of IFN-c as CD4 helper function has been implicated but needs more explorations in the future. Other mechanisms The CD40-independent CD4 T cell help has also been demonstrated. Pardoll et al. used both in vivo and in vitro CTL priming system to demonstrate that the CD4 T cell help in their model is CD40independent. They found the help signal is mediated by direct contact of CD4 T cell with the APC [17]. The nature of this contact
The major cellular subset responsible for IL-12 production in vivo is mature DC. It has been shown that IL-12 is a key regulator of CD8 CTL response in vitro and in vivo [21]. Thus it would be interesting to explore that whether IL-12 is the downstreamevent following DC maturation by CD40 ligation. Bianchi et al. reported that immunization of DC pulsed with a class I binding tumor peptide failed to elicit CD8 CTL response unless it was treated with anti-CD40 agonistic Ab before immunization. They further demonstrated that neutralization of IL-12 during CD40 activation in vitro can abrogates the adjuvant effect. The addition of exogenous IL-12 can overcome the requirement for anti-CD40 activation Ab [22,23]. Thus IL-12 might be a significant downstreamsignal for DC after CD40 ligation. However, using the classical OVA Ag, it also has been shown that DCs derived from IL-12 / mice pulsed with OVA peptide are fully capable of mounting a robust CD8 CTL response which is CD4 T cell and CD40L dependent [24]. Thus it seems that IL-12 is not required for the full activation of CD8 CTL in this CD4 T cell and CD40L dependent model. However, IL-12 neutralization or repletion was not used in this study. Also, the nature of Ag may decide different mechanisms. Further future studies need to link this discrepancy in different models. B7–CD28 signaling One important characteristic of DC maturation after CD40L activation is that DC can up-regulate the expression of the costimulatory molecules. Examining the costimulatory pathways might identify the pathway between DC and CD8 CTL which can finally lead to CTL activation. B7–CD28 pathway is one of the most important costimulatory pathways. Using a cross-priming model, it has been reported that both CD28 and B7-1/B7-2 are required for CD40-activated APC to cross prime CTL [25]. The priming by CD40 activated APC was prevented by blockade of CD28. However augmenting CD28 signals with an agonistic Ab can bypass the requirement for CD4 T cell help or CD40 activation. They also found that blockade of the negative regulatory B7 receptor CTLA-4 failed to prime CTL in the absence of CD4 T cell help. Thus this study supports the up-regulation of B7 molecules on APC leads to increased CD28 signaling and a commitment to cross-priming of CD4 T cell dependent CTL.
S. Zhang et al. / Biochemical and Biophysical Research Communications 384 (2009) 405–408
CD70–CD27 In recent years CD70 has been found to be another important costimulatory molecule expressed by mature DC. Bullock et al. recently reported that in their model they used MHC class II / DC pulsed with OVA peptide for immunization in order to avoid interaction with CD4 T cells in vivo. The CD4 T cell dependence of this DC immunization can be overcome with anti-CD40L agonistic Ab pretreatment [26]. The immune response elicited by DC is closely associated with CD70 expression on DC. They further found that the blockade of CD70 on DC with Ab substantially prevented the CD4 independence of the stimulated DC. They also found the interaction between CD70 on DC and CD27 on CD8 T cells needs to cooperate with B7–CD28 pathway. Thus they suggested that CD70–CD27 might be an important downstreamevent following CD4 T cell help through CD40–CD40L pathway. Arens et al. also reported that constitutive ligation of the tumor necrosis factor receptor family member CD27 by its ligand CD70 quantitatively and qualitatively augments CD8 CTL response to influenza virus infection and EL-4 tumor challenge in vivo [27]. Hendriks et al. further found that CD27 can promote the survival of activated T cells and CD27 can work cooperatively with CD28 in generation and establishment of effector CD8 CTL pool [28]. 4-1BBL–4-1BB Another TNFR member, 4-1BB, is expressed on CD8 T cells. It’s ligand, 4-1BBL, is expressed on DC. Diehl et al. reported that the agonistic anti-4-1BB Ab can effectively replace the need for CD4 T cell help in a cross priming model [29]. However, when they used blocking Ab against 4-1BBL which is expressed on DC, the CD8 CTL cannot be fully activated even in the presence of CD4 T cells. Furthermore, the agonistic anti-4-1BB Ab can convert an otherwise tolerogenic peptide vaccine into a formulation capable of efficient CTL priming. Blockade of the CD28 costimulatory pathway abrogates the capacity of agonistic anti-4-1BB Ab to trigger Th-independent CTL immunity. Thus they could show that 4-1BB mediated signal is positioned downstream of CD28 dependent costimulation of CD8 CTL. IL-15 IL-15, like IL-12, is made by DC after stimulation by CD4 helper T cells or by CD40L. A recent study showed that IL-15 could substitute for CD4 help to induce long-lived CD8 T cell memory and to prevent TRAIL-mediated apoptosis on secondary exposure to antigen, and also showed that IL-15 / DC could not fully mediate help even in the presence of normal helper T cells, implying that IL-15 was necessary as well as sufficient for help. Therefore, IL-15 may be a natural mediator of help [30].
The Ag specificity of CD4 T cell help The concept of CD4 T cell help of licensing of DC for CD8 CTL priming must be antigen specific has been long accepted. In different models, it has been shown that only when the Ag specific CD4 T cells are present, the optimal CD8 CTL can be elicited. Thus it has been concluded that Ag specific class II epitopes have to be included in the design of epitope-based vaccines [31–33]. Similarly even when class II epitope was not used to pulse DC in some cases, they still need Ag specific CD4 T cell help. In this case, the FBS additives used during the DC culture likely provide a source of MHC class II epitope to engage CD4 T cell help [24]. However, several recent studies are challenging this perception now. Sun et al. demonstrated recently that after immunization, when the helped effec-
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tor CD8 T cells (in the CD4 T cell sufficient environment at the time of immunization) were transferred into normal or class II deficient hosts, the CD8 T cells transferred into the CD4 deficient environment showed defective recall response [34]. However, the unhelped CD8 T cells (from the MHC class II deficient hosts) showed normal recall response when they were transferred into B6 mice compared with those transferred into MHC class II deficient mice. Thus the involvement of CD4 T cells in these experiments seems to be antigen nonspecific: recipient mice in the transfer experiments were unprimed. The CD4 T cell help in this case clearly is different from other models in which CD8 T cells and CD4 T cells need to cooperate for the response to the same Ag. It will be necessary in the future to determine whether these CD4 T cells are truly naïve cells or activated CD4 T cells to other undefined Ags. The timing of CD4 T cell help: at priming stage or later stage? Numerous studies have shown that CD4 depletion before or during priming generated functionally crippled CD8 T cell/memory cells. In these studies, depletion of CD4 T cells during boosting at later stage had little effect on memory CD8 T cell response [13,35,36]. However, recent studies conflict this theory. As described previously in this Nature Immunol study, the transfer of unhelped CD8 T cells into CD4 sufficient environment actually can restore the impaired memory CD8 T cell response [34]. In this case, the CD4 T cell help is in later stage but not the priming stage. Similarly, using the well characterized OVA system, several other independent studies also demonstrated CD4 T cell help can happen at later stage after priming [16,32]. More recently, a study showed that CD4 T cell help is necessary for the CD8 CTLs to avoid death by tumor necrosis factor related apoptosis inducing ligand. Regulation of Trail expression can therefore, account for the role of CD4 T cell help in the generation of CD8 T cell memory [1]. Thus there is still a debate about ‘‘programming”, or ‘‘maintenance” of CD8 T cells by CD4 T helper cells in this field. Concluding remarks The concept that CD4 T cell help is essential for CD8 CTL either for primary response or for secondary response has been well established. However how the CD4 T cell help can convert the DC to become fully competent for CD8 CTL activation, the timing and Ag specificity of CD4 T cell help, the detailed downstreamevents within DC and CD8 CTL after the CD40 ligation as well as the CD40-independent mechanisms need to be further extensively explored in the future. Acknowledgments Shanghai Pujiang Program 08PJ1407500, China (to S. Zhang); Research Grant 07B22 from First people’s hospital, Shanghai Jiaotong University, China (to S. Zhang); Chinese National Natural Science Foundation 30800279 (to J. Zhao). References [1] E.M. Janssen, N.M. Droin, E.E. Lemmens, M.J. Pinkoski, S.J. Bensinger, B.D. Ehst, et al., CD4+ T-cell help controls CD8+ T-cell memory via TRAIL-mediated activation-induced cell death, Nature 434 (2005) 88–93. [2] M.J. Bevan, Helping the CD8+ T-cell response, Nat. Rev. Immunol. 4 (2004) 595–602. [3] S.P. Schoenberger, R.E.M. Toes, E.I.H. van der Voort, R. Offringa, C.J.M. Melief, Tcell help for cytotoxic T lymphocytes is mediated by CD40–CD40L interactions, Nature 393 (1998) 480–483. [4] S.R.M. Bennett, F.R. Carbone, F. Karamalis, R.A. Flavell, J.F.A.P. Miller, W.R. Heath, Help for cytotoxic-T-cell responses is mediated by CD40 signalling, Nature 393 (1998) 478–480.
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[5] J.P. Ridge, F. Di Rosa, P. Matzinger, A conditioned dendritic cell can be a temporal bridge between a CD4+ T-helper and a T-killer cell, Nature 393 (1998) 474–478. [6] C. Bourgeois, B. Rocha, C. Tanchot, A role for CD40 expression on CD8+ T cells in the generation of CD8+ T cell memory, Science 297 (2002) 2060–2063. [7] B.O. Lee, L. Hartson, T.D. Randall, CD40-deficient, influenza-specific CD8 memory T cells develop and function normally in a CD40-sufficient environment, J. Exp. Med. 198 (2003) 1759–1764. [8] D.H. Schuurhuis, S. Laban, R.E.M. Toes, P. Ricciardi-Castagnoli, M.J. Kleijmeer, E.I.H. van der Voort, et al., Immature dendritic cells acquire CD8+ cytotoxic T lymphocyte priming capacity upon activation by T helper cell-independent or dependent stimuli, J. Exp. Med. 192 (2000) 145–150. [9] C.M. Smith, N.S. Wilson, J. Waithman, J.A. Villadangos, F.R. Carbone, W.R. Heath, et al., Cognate CD4+ T cell licensing of dendritic cells in CD8+ T cell immunity, Nat. Immunol. 5 (2004) 1143–1148. [10] J.A. Keene, J. Forman, Helper activity is required for the in vivo generation of cytotoxic T lymphocytes, J. Exp. Med. 155 (1982) 768–782. [11] J. Steiger, P.W. Nickerson, W. Steurer, M. Moscovitch-Lopatin, T.B. Strom, IL-2 knockout recipient mice reject islet cell allografts, J. Immunol. 155 (1995) 489–498. [12] J.E. Buhlmann, M. Gonzalez, B. Ginther, A. Panoskaltsis-Mortari, B.R. Blazar, D.L. Greiner, et al., Cutting edge: sustained expansion of CD8+ T cells requires CD154 expression by Th cells in acute graft versus host disease, J. Immunol. 162 (1999) 4373–4376. [13] E.M. Janssen, E.E. Lemmens, T. Wolfe, U. Christen, M.G. von Herrath, S.P. Schoenberger, CD4+ T cells are required for secondary expansion and memory in CD8+ T lymphocytes, Nature 421 (2003) 852–856. [14] M.A. Williams, A.J. Tyznik, M.J. Bevan, Interleukin-2 signals during priming are required for secondary expansion of CD8+ memory T cells, Nature 441 (2006) 890–893. [15] C.R. Ferrone, M.-A. Perales, S.M. Goldberg, C.J. Somberg, D. HirschhornCymerman, P.D. Gregor, et al., Adjuvanticity of plasmid DNA encoding cytokines fused to immunoglobulin Fc domains, Clin. Cancer Res. 12 (2006) 5511–5519. [16] U. Kumaraguru, K. Banerjee, B.T. Rouse, In vivo rescue of defective memory CD8+ T cells by cognate helper T cells, J. Leukoc Biol. 78 (2005) 879–887. [17] Z. Lu, L. Yuan, X. Zhou, E. Sotomayor, H.I. Levitsky, D.M. Pardoll, CD40independent pathways of T cell help for priming of CD8+ cytotoxic T lymphocytes, J. Exp. Med. 191 (2000) 541–550. [18] R. Josien, H.-L. Li, E. Ingulli, S. Sarma, B.R. Wong, M. Vologodskaia, et al., TRANCE, a tumor necrosis factor family member, enhances the longevity and adjuvant properties of dendritic cells in vivo, J. Exp. Med. 191 (2000) 495–502. [19] E. Allison Green, R.A. Flavell, TRANCE-RANK, a new signal pathway involved in lymphocyte development and T cell activation, J. Exp. Med. 189 (1999) 1017– 1020. [20] M.F. Bachmann, B.R. Wong, R. Josien, R.M. Steinman, A. Oxenius, Y. Choi, TRANCE, a tumor necrosis factor family member critical for CD40 ligand-independent T helper cell activation, J. Exp. Med. 189 (1999) 1025–1031.
[21] J.M. Curtsinger, D.C. Lins, M.F. Mescher, Signal 3 determines tolerance versus full activation of naive CD8 T cells: dissociating proliferation and development of effector function, J. Exp. Med. 197 (2003) 1141–1151. [22] R. Bianchi, U. Grohmann, C. Vacca, M.L. Belladonna, M.C. Fioretti, P. Puccetti, Autocrine IL-12 is involved in dendritic cell modulation via CD40 ligation, J. Immunol. 163 (1999) 2517–2521. [23] K.K. Per Grufman, Innate and adaptive immunity to tumors: IL-12 is required for optimal responses, Eur. J. Immunol. 30 (2000) 1088–1093. [24] Y. Wan, L. Lu, J.L. Bramson, S. Baral, Q. Zhu, A. Pilon, et al., Dendritic cellderived IL-12 is not required for the generation of cytotoxic, IFN-{gamma}secreting, CD8+ CTL in vivo, J. Immunol. 167 (2001) 5027–5033. [25] K.R. Prilliman, E.E. Lemmens, G. Palioungas, T.G. Wolfe, J.P. Allison, A.H. Sharpe, et al., Cutting edge: a crucial role for B7–CD28 in transmitting T help from APC to CTL, J. Immunol. 169 (2002) 4094–4097. [26] T.N.J. Bullock, H. Yagita, Induction of CD70 on dendritic cells through CD40 or TLR stimulation contributes to the development of CD8+ T cell responses in the absence of CD4+ T Cells, J. Immunol. 174 (2005) 710–717. [27] R. Arens, K. Schepers, M.A. Nolte, M.F. van Oosterwijk, R.A.W. van Lier, T.N.M. Schumacher, et al., Tumor rejection induced by CD70-mediated quantitative and qualitative effects on effector CD8+ T cell formation, J. Exp. Med. 199 (2004) 1595–1605. [28] J. Hendriks, Y. Xiao, J. Borst, CD27 promotes survival of activated T cells and complements CD28 in generation and establishment of the effector T cell pool, J. Exp. Med. 198 (2003) 1369–1380. [29] L. Diehl, G.J.D. van Mierlo, A.T. den Boer, E. van der Voort, M. Fransen, L. van Bostelen, et al., In vivo triggering through 4-1BB enables Th-independent priming of CTL in the presence of an intact CD28 costimulatory pathway, J. Immunol. 168 (2002) 3755–3762. [30] S. Oh, L.P. Perera, M. Terabe, L. Ni, T.A. Waldmann, J.A. Berzofsky, IL-15 as a mediator of CD4+ help for CD8+ T cell longevity and avoidance of TRAILmediated apoptosis, Proc. Nat. Acad. Sci. USA 105 (2008) 5201–5206. [31] F. Ossendorp, E. Mengede, M. Camps, R. Filius, C.J.M. Melief, Specific T helper cell requirement for optimal induction of cytotoxic T lymphocytes against major histocompatibility complex class II negative tumors, J. Exp. Med. 187 (1998) 693–702. [32] F.G. Gao, V. Khammanivong, W.J. Liu, G.R. Leggatt, I.H. Frazer, G.J.P. Fernando, Antigen-specific CD4+ T-cell help is required to activate a memory CD8+ T cell to a fully functional tumor killer cell, Cancer Res. 62 (2002) 6438–6441. [33] M. Shirai, C.D. Pendleton, J. Ahlers, T. Takeshita, M. Newman, J.A. Berzofsky, Helper-cytotoxic T lymphocyte (CTL) determinant linkage required for priming of anti-HIV CD8+ CTL in vivo with peptide vaccine constructs, J. Immunol. 152 (1994) 549–556. [34] J.C. Sun, M.A. Williams, M.J. Bevan, CD4+ T cells are required for the maintenance, not programming, of memory CD8+ T cells after acute infection, Nat. Immunol. 5 (2004) 927–933. [35] D.J. Shedlock, H. Shen, Requirement for CD4 T cell help in generating functional CD8 T cell memory, Science 300 (2003) 337–339. [36] J.C. Sun, M.J. Bevan, Defective CD8 T cell memory following acute infection without CD4 T cell help, Science 300 (2003) 339–342.
Contents lists available at ScienceDirect
Biochemical and Biophysical Research Communications journal homepage: www.elsevier.com/locate/ybbrc
Mini Review
The role of CD4 T cell help for CD8 CTL activation Sheng Zhang a,d,*,1, Hongjun Zhang b,1, Jiandong Zhao c a
Medical Oncology, First People’s Hospital, Shanghai Jiaotong University, 650 Xinsongjiang Road, Shanghai 201620, China Medical Oncology, The Medical School Hospital of Qingdao University, Qingdao, China Department of Radiation Oncology, Cancer Hospital of Fudan University, Shanghai, China d Shanghai Key Laboratory of Pancreatic Diseases, Shanghai, China b c
a r t i c l e
i n f o
Article history: Received 19 April 2009 Available online 3 May 2009
a b s t r a c t CD4 T cells play an important role in the initiation and persistence of CD8 T cells responses. In this review, we report on and evaluate the mechanisms by which CD4 T cells contribute to activation of CD8 T cells and the signal pathways of the down-streaming events after CD4 T cell help. Ó 2009 Elsevier Inc. All rights reserved.
Keywords: CD4 T cells CD8 T cells Signal pathway
CD8 CTL cells are essential for protection against viruses, intracellular bacteria infection and tumor cells. Understanding the mechanism for CTL activation, survival and long term maintenance has become one focus for many immunologists. Numerous studies in recent years have reached a consensus that CD4 T cell help is essential for CD8 CTL priming. The help provided by CD4 T lymphocytes during the priming of CD8 T lymphocytes confers a key feature of immune memory: the capacity for autonomous secondary expansion following re-encounter with antigen. Once primed in the presence of CD4 T cells, helped CD8 T cells acquire the ability to undergo a second round of clonal expansion upon restimulation in the absence of T cell help. The CD8 T cells that are not helped by CD4 T cells, in contrast, can mediate effector functions such a s cytotoxicity and cytokine secretion upon restimulation, but do not undergo a second round of clonal expansion [1,2]. These disparate responses have features of being programmed, that is, guided by signals that are transmitted to naïve CD8 T cells during priming, which encode specific fates for their clonal progeny. In this regard, exploring the instructional programme that governs the secondary response of CD8 T cells has significant impact on the future design of CD8 CTL based vaccination. This paper will discuss the mechanisms of CD4 T cell help, the DC licensing model and the downstreamevents following CD4 T cell help pathway.
* Corresponding author. Fax: +86 533 3177425. E-mail address: [email protected] (S. Zhang). 1 These authors contributed equally to this study. 0006-291X/$ - see front matter Ó 2009 Elsevier Inc. All rights reserved. doi:10.1016/j.bbrc.2009.04.134
The mechanisms of CD4 T cell help CD40–CD40L signaling pathway Recent findings have established a DC licensing model. In this model, the CD40L expressing CD4 T cells can interact with the professional APC-DC through the CD40–CD40L pathway. This interaction can lead to the activation/licensing of DC, then the licensed DC can activate CD8 CTL cells. This model was originally proposed/ established by 3 papers in Nature in 1998 [3–5]. In those studies, they found that CTL response was dependent on both CD40 and CD40 ligand and that in the absence of CD4 T helper cells an agonistic antibody to CD40 can substitute for CD4 T cell help. Since the APC was the only known cell to express CD40 and CD4 T helper cells were known to express CD40 ligand upon activation, it was concluded that CD40-dependent licensing of APC was the crucial nature of CD4 T cell help. This proposed licensing model was seriously challenged by another study recently in Science [6]. In this study, the authors could show that CD4 T cell help for the differentiation of CD8 T cells into memory T cells can occur even when all APCs are CD40-deficient. They showed that CD8 T cells can express CD40 transiently after activation. Moreover, CD40-deficient CD8 T cells can never differentiate into memory cells, even in the presence of CD4 T helper cells. They cannot receive helper signals from antigen specific CD4 T cells. Thus, as B cells, the helper signals required for differentiation of CD8 T cells into memory cells appear to pass through the CD40 molecule expressed by activated CD8 T cells. In this model, although CD4 T cell help is CD40-dependent, it is through a direct contact between CD4 T cells and CD8 T cells. The role of DC in this case is to provide the chance that CD4 T cells
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and CD8 T cells can be put into proximity for their contact. Thus this study gave direct challenge to the proposed licensing model. Lee and colleagues demonstrated that optimal CD8 CTL response to influenza is dependent on CD40 signaling. However, both primary and secondary response to influenza require CD40 expression on non-T cells [7]. Thus in this case, CD4 T helper cells do not activate CD8 T cells directly through CD40 signaling. With this study, they could show that the direct interaction between CD4 T cells and CD8 T cells through CD40–CD40L signaling is not a universal mechanism for CD4 T cell help. Schuurhuis and colleagues further explored this proposed model. They showed that the DC cell line D1 could activate CD8 CTL with the help of CD4 Th cell [8]. However, when the D1 cells were stimulated with agonistic antiCD40 Ab, they can activate CD8 CTL in a CD4 T cell-independent way, thus further suggesting this DC license model might be true. In 2004, a Nature Immunol paper further gave direct in vivo evidence for DC licensing by Th cells [9]. In this study, they could show that the in vivo sorted DC, only when they were licensed by CD4 T cells, could mediate a memory T cell response. Thus, this model of licensing of DC by CD4 Th cells for CTL priming was finally firmly established.
mediated signal was not identified. It has been reported that signaling of osteoprotgerin ligand (OPGL), another TNF-receptor family member expressed by DC has some similar effects to CD40 signaling. In other words, CD4 T cells can interact with DC through the TRANCE (tumor necrosis factor-related-activation-induced cytokine) pathway which finally leads to the activation of DC [18,19]. Also the CD4 T cells can acquire the effector function for virus protection in this case. Thus TRANCE has been documented as a signaling pathway leading to CD4 T cell activation independent of CD40L [20]. But the use of this signaling pathway by helper cells involved in CTL induction has not been reported. It’s tempting in the future to explore whether this pathway is also used by CD4 T cells for CD8 CTL priming independently of CD40 pathway. The downstreamevents of CD4 T cell help: what happens to DC and CD8 T cells following CD40 ligation? So far, the most characterized and accepted model is still the CD4 Th cells, DC, CD8 CTL model and CD40–CD40L pathway is the major signaling pathway between CD4 Th cells and DC. One important problem would be to address what happened to DC after CD4 T cell licensing so that DC can activate CTL?
IL-2 IL-12 One important characteristic of CD4 T cells is that they can produce multiple cytokines. Thus cytokines have lone been suggested that they can be the mediators of CD4 T cell help. In vitro studies have found that IL-2 is essential for the effective generation of CTL response when helper T cells and CTL were combined in the culture flask [10]. However, conflicting results exist since CTL response to viruses can be generated in IL-2 knock out mice [11]. More recently, using a graft versus host disease model, it was found that the ability of CD4 T helper cells to produce IL-2 was essential for sustained expansion of alloreactive CTL [12]. Using a cross-priming model, several recent studies have demonstrated that the absence of CD4 T cell help for CD8 CTL priming can be overcome by the provision of exogenous IL-2, thus establishing the role of IL-2 for CD4 T cell help [13,14]). More studies need to be done in order to confirm the role of IL-2 as the helper of CD8 CTL or this role can only exist in some certain cases. IFN-c IFN-c is one of the most important type 1 cytokines. It has been shown that IFN-c can enhance the cytotoxicity of CTL in addition to being one of the important effector arms for CD8 CTL. In a recent report, Wolchok and colleagues demonstrated that in a DNA vaccination model, the CTL is fully dependent on CD4 T cell help. However, this CD4 T cell-dependent CTL is lost in IFN-c / mice. It can be restored when recombinant IFN-c was injected in vivo at the priming stage [15], suggesting IFN-c is the essential mediator for CD4 T cell help in this model. A more recent study by Kumaraguru et al. used the well characterized HSV and OVA model system. They found that the helper function of CD4 T cells is mainly mediated by IFN-c producing CD4 Th cells but not IL-2 producing CD4 Th cells [16]. Thus the role of IFN-c as CD4 helper function has been implicated but needs more explorations in the future. Other mechanisms The CD40-independent CD4 T cell help has also been demonstrated. Pardoll et al. used both in vivo and in vitro CTL priming system to demonstrate that the CD4 T cell help in their model is CD40independent. They found the help signal is mediated by direct contact of CD4 T cell with the APC [17]. The nature of this contact
The major cellular subset responsible for IL-12 production in vivo is mature DC. It has been shown that IL-12 is a key regulator of CD8 CTL response in vitro and in vivo [21]. Thus it would be interesting to explore that whether IL-12 is the downstreamevent following DC maturation by CD40 ligation. Bianchi et al. reported that immunization of DC pulsed with a class I binding tumor peptide failed to elicit CD8 CTL response unless it was treated with anti-CD40 agonistic Ab before immunization. They further demonstrated that neutralization of IL-12 during CD40 activation in vitro can abrogates the adjuvant effect. The addition of exogenous IL-12 can overcome the requirement for anti-CD40 activation Ab [22,23]. Thus IL-12 might be a significant downstreamsignal for DC after CD40 ligation. However, using the classical OVA Ag, it also has been shown that DCs derived from IL-12 / mice pulsed with OVA peptide are fully capable of mounting a robust CD8 CTL response which is CD4 T cell and CD40L dependent [24]. Thus it seems that IL-12 is not required for the full activation of CD8 CTL in this CD4 T cell and CD40L dependent model. However, IL-12 neutralization or repletion was not used in this study. Also, the nature of Ag may decide different mechanisms. Further future studies need to link this discrepancy in different models. B7–CD28 signaling One important characteristic of DC maturation after CD40L activation is that DC can up-regulate the expression of the costimulatory molecules. Examining the costimulatory pathways might identify the pathway between DC and CD8 CTL which can finally lead to CTL activation. B7–CD28 pathway is one of the most important costimulatory pathways. Using a cross-priming model, it has been reported that both CD28 and B7-1/B7-2 are required for CD40-activated APC to cross prime CTL [25]. The priming by CD40 activated APC was prevented by blockade of CD28. However augmenting CD28 signals with an agonistic Ab can bypass the requirement for CD4 T cell help or CD40 activation. They also found that blockade of the negative regulatory B7 receptor CTLA-4 failed to prime CTL in the absence of CD4 T cell help. Thus this study supports the up-regulation of B7 molecules on APC leads to increased CD28 signaling and a commitment to cross-priming of CD4 T cell dependent CTL.
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CD70–CD27 In recent years CD70 has been found to be another important costimulatory molecule expressed by mature DC. Bullock et al. recently reported that in their model they used MHC class II / DC pulsed with OVA peptide for immunization in order to avoid interaction with CD4 T cells in vivo. The CD4 T cell dependence of this DC immunization can be overcome with anti-CD40L agonistic Ab pretreatment [26]. The immune response elicited by DC is closely associated with CD70 expression on DC. They further found that the blockade of CD70 on DC with Ab substantially prevented the CD4 independence of the stimulated DC. They also found the interaction between CD70 on DC and CD27 on CD8 T cells needs to cooperate with B7–CD28 pathway. Thus they suggested that CD70–CD27 might be an important downstreamevent following CD4 T cell help through CD40–CD40L pathway. Arens et al. also reported that constitutive ligation of the tumor necrosis factor receptor family member CD27 by its ligand CD70 quantitatively and qualitatively augments CD8 CTL response to influenza virus infection and EL-4 tumor challenge in vivo [27]. Hendriks et al. further found that CD27 can promote the survival of activated T cells and CD27 can work cooperatively with CD28 in generation and establishment of effector CD8 CTL pool [28]. 4-1BBL–4-1BB Another TNFR member, 4-1BB, is expressed on CD8 T cells. It’s ligand, 4-1BBL, is expressed on DC. Diehl et al. reported that the agonistic anti-4-1BB Ab can effectively replace the need for CD4 T cell help in a cross priming model [29]. However, when they used blocking Ab against 4-1BBL which is expressed on DC, the CD8 CTL cannot be fully activated even in the presence of CD4 T cells. Furthermore, the agonistic anti-4-1BB Ab can convert an otherwise tolerogenic peptide vaccine into a formulation capable of efficient CTL priming. Blockade of the CD28 costimulatory pathway abrogates the capacity of agonistic anti-4-1BB Ab to trigger Th-independent CTL immunity. Thus they could show that 4-1BB mediated signal is positioned downstream of CD28 dependent costimulation of CD8 CTL. IL-15 IL-15, like IL-12, is made by DC after stimulation by CD4 helper T cells or by CD40L. A recent study showed that IL-15 could substitute for CD4 help to induce long-lived CD8 T cell memory and to prevent TRAIL-mediated apoptosis on secondary exposure to antigen, and also showed that IL-15 / DC could not fully mediate help even in the presence of normal helper T cells, implying that IL-15 was necessary as well as sufficient for help. Therefore, IL-15 may be a natural mediator of help [30].
The Ag specificity of CD4 T cell help The concept of CD4 T cell help of licensing of DC for CD8 CTL priming must be antigen specific has been long accepted. In different models, it has been shown that only when the Ag specific CD4 T cells are present, the optimal CD8 CTL can be elicited. Thus it has been concluded that Ag specific class II epitopes have to be included in the design of epitope-based vaccines [31–33]. Similarly even when class II epitope was not used to pulse DC in some cases, they still need Ag specific CD4 T cell help. In this case, the FBS additives used during the DC culture likely provide a source of MHC class II epitope to engage CD4 T cell help [24]. However, several recent studies are challenging this perception now. Sun et al. demonstrated recently that after immunization, when the helped effec-
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tor CD8 T cells (in the CD4 T cell sufficient environment at the time of immunization) were transferred into normal or class II deficient hosts, the CD8 T cells transferred into the CD4 deficient environment showed defective recall response [34]. However, the unhelped CD8 T cells (from the MHC class II deficient hosts) showed normal recall response when they were transferred into B6 mice compared with those transferred into MHC class II deficient mice. Thus the involvement of CD4 T cells in these experiments seems to be antigen nonspecific: recipient mice in the transfer experiments were unprimed. The CD4 T cell help in this case clearly is different from other models in which CD8 T cells and CD4 T cells need to cooperate for the response to the same Ag. It will be necessary in the future to determine whether these CD4 T cells are truly naïve cells or activated CD4 T cells to other undefined Ags. The timing of CD4 T cell help: at priming stage or later stage? Numerous studies have shown that CD4 depletion before or during priming generated functionally crippled CD8 T cell/memory cells. In these studies, depletion of CD4 T cells during boosting at later stage had little effect on memory CD8 T cell response [13,35,36]. However, recent studies conflict this theory. As described previously in this Nature Immunol study, the transfer of unhelped CD8 T cells into CD4 sufficient environment actually can restore the impaired memory CD8 T cell response [34]. In this case, the CD4 T cell help is in later stage but not the priming stage. Similarly, using the well characterized OVA system, several other independent studies also demonstrated CD4 T cell help can happen at later stage after priming [16,32]. More recently, a study showed that CD4 T cell help is necessary for the CD8 CTLs to avoid death by tumor necrosis factor related apoptosis inducing ligand. Regulation of Trail expression can therefore, account for the role of CD4 T cell help in the generation of CD8 T cell memory [1]. Thus there is still a debate about ‘‘programming”, or ‘‘maintenance” of CD8 T cells by CD4 T helper cells in this field. Concluding remarks The concept that CD4 T cell help is essential for CD8 CTL either for primary response or for secondary response has been well established. However how the CD4 T cell help can convert the DC to become fully competent for CD8 CTL activation, the timing and Ag specificity of CD4 T cell help, the detailed downstreamevents within DC and CD8 CTL after the CD40 ligation as well as the CD40-independent mechanisms need to be further extensively explored in the future. Acknowledgments Shanghai Pujiang Program 08PJ1407500, China (to S. Zhang); Research Grant 07B22 from First people’s hospital, Shanghai Jiaotong University, China (to S. Zhang); Chinese National Natural Science Foundation 30800279 (to J. Zhao). References [1] E.M. Janssen, N.M. Droin, E.E. Lemmens, M.J. Pinkoski, S.J. Bensinger, B.D. Ehst, et al., CD4+ T-cell help controls CD8+ T-cell memory via TRAIL-mediated activation-induced cell death, Nature 434 (2005) 88–93. [2] M.J. Bevan, Helping the CD8+ T-cell response, Nat. Rev. Immunol. 4 (2004) 595–602. [3] S.P. Schoenberger, R.E.M. Toes, E.I.H. van der Voort, R. Offringa, C.J.M. Melief, Tcell help for cytotoxic T lymphocytes is mediated by CD40–CD40L interactions, Nature 393 (1998) 480–483. [4] S.R.M. Bennett, F.R. Carbone, F. Karamalis, R.A. Flavell, J.F.A.P. Miller, W.R. Heath, Help for cytotoxic-T-cell responses is mediated by CD40 signalling, Nature 393 (1998) 478–480.
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