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Autoreactive effector CD4+ T cells have been associated with the pathogenesis of autoimmune disorders. Early studies implicated the interferon (IFN)-γ-producing T helper (Th)1 subset of CD4+ cells as the causal agents in the pathogenesis of autoimmunity. However, further studies have suggested a more complex story. In models thought to be driven by Th1 cells, mice lacking the hallmark Th1 cytokine IFN-γ were not protected but tended to have enhanced susceptibility to disease. Identification of the IL-17-producing CD4+ effector cell lineage (Th17) has helped shed light on this issue. Th17 effector cells are induced in parallel to Th1, and, like Th1, polarized Th17 cells have the capacity to cause inflammation and autoimmune disease. This, together with the finding that deficiency of the Th17-related cytokine IL-23 but not the Th1-related cytokine IL-12 causes resistance, led to the notion that Th17 cells are the chief contributors to autoimmune tissue inflammation. Nevertheless, mice lacking IL-17 are not protected from disease and display elevated numbers of IFN-γ-producing CD4+ T cells, and, in some cases, lack of IFN-γ does confer resistance. Recent studies report overlapping as well as differential roles of these cells in tissue inflammation, which suggests the existence of a more complex relationship between these two effector T-cell subsets than has hitherto been suspected. This review will attempt to bring together current information regarding interaction, balance, and collaborative potential between the Th1 and Th17 effector lineages.
Autoreactive effector CD4+ T cells have been associated with the pathogenesis of several autoimmune disorders, including multiple sclerosis, Crohn’s disease, and rheumatoid arthritis. Exactly 20 years ago, Mosmann and colleagues introduced the paradigmshifting concept that effector CD4+ T cells were subdivided into two subsets, T helper (Th)1 and Th2, on the basis of their functions and signature cytokine profiles. Th2 cells, which produce the cytokines IL-4, IL-5, and IL-13, are involved in allergic responses and the clearance of extracellular pathogens, such as worms.1 IFN-γ-producing Th1 cells, on the other hand, contribute to the elimination of intracellular pathogens and are involved in cell-mediated and delayed-type hypersensitivity responses.1 In earlier studies, Th1 cells were implicated as the causal agents in the pathogenesis of autoimmunity as these cells possessed the ability to transfer disease and elevated levels of IFN-γ were detected in vivo in areas of inflammation.
However, studies in several autoimmune disease models suggested a more complex story. Mice genetically deficient in IFN-γ not only were not protected but also exhibited enhanced susceptibility in many models of autoimmunity.2–4 The identification of Th17 cells, a CD4+ T-cell subset that produces IL-17, has helped to shed some light on this apparent paradox. These cells, like Th1 cells, have the capacity to cause T-cell-mediated inflammation and autoimmune disease. Moreover, the Th17 response is remarkably elevated in mice lacking IFN-γ. However, we have recently reported that in experimental autoimmune uveitis (EAU), a model for human posterior uveitis, mice lacking IL-17 are also not protected and display elevated numbers of IFN-γ-producing CD4+ T cells.5,6 This raises the questions, what is the relationship between Th1 and Th17 cells? Might they cooperate with one another to propagate the inflammatory response? Do they merely play a role in regulating one another? This review will focus on how interactions between Th1 and Th17 responses may contribute to autoimmunity.
While the primary function of both Th1 and Th2 cells is in host defense, much attention has been focused on their role(s) in autoimmune disease. The traditional model suggests that in tissue-specific autoimmune diseases, such as EAU and experimental autoimmune encephalomyelitis (EAE), a model for human multiple sclerosis (MS), Th1 cells are the pro-inflammatory effectors of autoimmunity, whereas Th2 cells function to antagonize this effect.7,8 In early studies there were several lines of evidence to support this notion. In mice susceptible to EAE, interferon (IFN)-γ levels within the central nervous system (CNS) correlate with disease severity, with high levels detected at the peak of disease that fall as the disease spontaneously subsides into remission.9 Infiltrating CD4+ T cells were identified as the source of this IFN-γ,10 and adoptive transfer of IFN-γ-producing T-cell lines has been shown to induce EAE, EAU, and other autoimmune pathologies.11 The role of Th1 in EAE was further supported by reports that mice lacking the Th1 lineage-specific transcription factors, T-bet and signal transducer and activator of transcription 4 (STAT4), are protected from disease. Importantly, administration of the Th1 cytokine, IFN-γ, to human MS patients based on its apparent (paradoxical) protective effect in animal models turned out to exacerbate clinical symptoms.12 Several investigators reported, what appeared to be emphatic evidence for the critical role of the Th1 response, that mice lacking the IL-12p40 subunit were protected in a variety of autoimmune models.1,13–15 This critical finding in favor of the Th1 response was subsequently demonstrated to be a result of the lack of IL-23, not IL-12,16 as discussed later in this review. Data consistent with the notion that Th1 cells were associated with pathology were also reported in other models of autoimmunity, including EAU17,18 and collagen-induced arthritis (CIA), a mouse model for human rheumatoid arthritis.19
Although a preponderance of evidence in the 1990s and early 2000s pointed to Th1 cells as mediators of autoimmune inflammation, some studies even at that time contradicted this notion. As previously mentioned, targeting IFN-γ either by neutralizing antibody or by genetic deletion in mice does not confer resistance to EAE, CIA, or EAU.2–4,20,21 In fact, in all cases, the mice developed exacerbated symptoms. In addition to this, findings in mice lacking IL-12p35, the IL-12 subunit with which IL-12p40 combines, contradicted the findings in p40-deficient mice. IL-12p35-deficient mice exhibited a phenotype similar to IFN-γ-deficient mice, with exacerbated disease symptoms in EAE and EAU.5,22,23
These paradoxical results imply that IFN-γ, considered to be a defining cytokine of the Th1 response, may actually play a protective role in autoimmunity. Even more intriguing, classical Th2 responses (IL-4) were not found to be elevated in these studies,3,22,23 raising the possibility that a disease-mediating Th subset other than the Th2 lineage might be at play in IFN-γ-deficient and p35-deficient animals.
The identification of the Th17 subset of effector CD4+ T cells has provided a new understanding as to the underlying mechanisms of autoimmunity. This recently described Th cell lineage is a potent producer of IL-17A and IL-17F, both of which belong to the IL-17 family of cytokines that includes IL-17B, IL-17C, IL-17D, and IL-17E (also known as IL-25).24 In terms of function, IL-17A and IL-17F target various (mostly nonlymphoid) cell types, including fibroblasts, endothelial cells, epithelial cells, keratinocytes, and macrophages, and induces the production of a milieu of cytokines (such as IL-6, granulocyte colony-stimulating factor [G-CSF], granulocyte–macrophage CSF [GM-CSF], IL-1, transforming growth factor [TGF]-β , tumor necrosis factor [TNF]-α), chemokines (including monocyte chemoattractant protein [MCP]-1, cytokine-induced neutrophil chemoattractant [KC], and macrophage inflammatory protein [MIP]-2), and prostaglandins (e.g., PGE2).16,25–28 An important outcome of these effects is the stimulation and attraction of neutrophils to the site of inflammation.5,25 IL-17 has also been shown to play a role in the production of matrix metalloproteinase (MMP) that functions to degrade tissue during an inflammatory response.25
In addition to IL-17A and F, Th17 cells produce IL-6, TNF-α, IL-21, and IL-22, whose role has been somewhat controversial.25,29,30 IL-22 belongs to the IL-10 family of cytokines and is produced, in large part, by CD4+ T cells and natural killer (NK) cells.30 IL-22 signals through a heterodimeric receptor composed of IL-22R1 and IL-10Rβ, and while IL-10Rβ is ubiquitously expressed, IL-22R1 expression is restricted entirely to cells of nonlymphoid origin, such as epithelial cells and fibroblasts.30,31 In this way, IL-22 is thought to exert its effects directly on the tissue rather than on lymphoid effector cells and has been shown to be particularly important in mucosal host defense in the gut and lung.32,33
In parallel to Th1 and Th2 cells, Th17 cells have their own distinct set of differentiation factors. It was first thought that this role of differentiation was played by IL-23 as IL-23 was found to induce Th17 proliferation in vitro16,34 and appeared analogous to the role of the closely related IL-12 for the Th1 lineage. IL-23 is a member of the IL-12 family of cytokines and shares a common p40 subunit with IL-12 but incorporates a unique p19 subunit in place of IL-12p35.35 However, further investigation revealed that IL-23 could only drive the proliferation of memory T cells that were already expressing IL-23R but was ineffective in inducing IL-17 production from naive cells.26,36 Rather, it is the combined effect of IL-6 and TGF-β that drives the initial commitment of naive T cells to the Th17 phenotype, although continued IL-23 signaling is essential for their survival and maintenance.26,36,37 This discovery unexpectedly revealed a reciprocal developmental pathway shared by Th17 and regulatory T cells as the presence of IL-6 completely inhibits the upregulation of Foxp3 and the acquisition of a suppressive phenotype that otherwise occurs when naive T cells are activated in the presence of TGF-β alone.37,38 Interestingly, another Th17-produced cytokine, IL-21, which is a member of the IL-2 family of cytokines, can amplify the Th17 response in an autocrine manner and even substitute for IL-6 in the induction of Th17.29 However, unlike IL-6, two recent studies have confirmed that IL-21 signaling enhances rather than inhibits the suppressive function of regulatory T cells, adding another layer to the complexity.39,40 In the aggregate, these findings demonstrate a complex interplay of cytokines that influence the differentiation and propagation of the Th17 response that is intertwined with the induction and function of Foxp3+ regulatory T cells.
Just as Th17 cells require a set of differentiation cytokines distinct from those required for Th1 and Th2 differentiation, Th17 cells also require distinct transcription factors and signaling molecules. Th17 cells do not express either T-bet or GATA-3, the transcription factors required for both Th1 and Th2 differentiation, respectively.1,41 Instead, Th17 cells express the orphan nuclear receptor RORγt, and mice with T cells deficient in this transcription factor fail to develop Th17 cells.42 RORγt expression is regulated by STAT3 signaling,43 which occurs downstream of T-cell activation through the T-cell receptor (TCR), TGF-β, and IL-6 receptors. However, recent studies have implicated another prototypic Th2 transcription factor, c-Maf, in the differentiation of Th17 cells, contributing to blurring of the sharp distinction between Th subsets.44
Because Th17 cells are associated with the production of inflammatory mediators, much attention has been given to their potentially pathogenic role in autoimmune disease. Supporting such a role are reports that IL-17 expression is increased at the site of inflammation in patients with rheumatoid arthritis, psoriasis, MS, and uveitis.45–50
The role of the Th17 response in autoimmunity has been examined experimentally within the context of various animal models. Mice genetically deficient in IL-17A were reported to be less susceptible to CIA and EAE,51 and neutralization of IL-17 by treatment with anti-IL-17 antibodies in vivo reduces severity of EAE52 and protects mice from EAU.5 The loss of Th17 regulatory components, such as RORγt and IL-6, have also been shown to result in attenuated autoimmune disease and in lack of tissueinfiltrating Th17 cells.42
In view of the role of IL-23 in maintenance of the Th17 response, various studies have examined the role of this cytokine in autoimmune disease. Mice genetically deficient in p19, the unique subunit of IL-23, are resistant to EAU, EAE, and CIA.5,15,53 Importantly, this outcome explains the inconsistency previously reported in mice lacking IL-12p40 compared with p35. It is now understood that the protection by deletion of the IL-12p40 chain, shared between IL-23 and IL-12, can be attributed to the absence of IL-23 and Th17, not IL-12 and Th1. These findings led some investigators to propose that it is Th17 cells that drive pathology of tissue-specific autoimmune disease, whereas Th1 cells have a minor role, if any.15,16,49
The role of Th1 in autoimmune disease, however, cannot be discounted. In the EAU model, mice genetically deficient in IL-17 still develop disease,5 and T cells polarized to the Th1 phenotype are able to transfer EAU and EAE with comparable severity to Th17-polarized cells.5,54 T-bet-deficient mice that lack a Th1 response are resistant to EAE despite the fact that T-bet−/− mice have no defect in generation of Th17 cells.41,55–58 Finally, in a nonclassical model of EAU in which peptide-pulsed dendritic cells rather than active immunization with interphotoreceptor retinoid-binding protein in Complete Freund’s Adjuvant (CFA) were used to induce disease, the ability to generate IFN-γ-producing Th1 effector cells was found to be essential for EAU pathogenesis. Similarly to T-bet-deficient mice, IFN-γ-deficient mice in this model were protected from disease, despite elevated levels of IL-17.5,59
Why do Th17 cells appear to drive pathology in some models and Th1 cells do so in others, even for the same autoimmune disease? And as a corollary, what determines the dominant effector phenotype? Notably, the autoimmunity models that demonstrate a Th17-dependent pathology in which IFN-γ plays a protective role are all induced by immunization with autoantigen emulsified in CFA. We suggest that the conditions under which the initial exposure to antigen occurs, including the type of innate receptors that are stimulated and the type of antigen-presenting cells (APCs) participating, affects the effector dominance in the subsequent adaptive autoimmune response (Fig. 1). The heat-killed mycobacteria in CFA activate a variety of APCs in the draining lymph node through innate immunity receptors, including Toll-like receptors (TLR) and NOD-like receptors (NLR), resulting in a Th17-dominated response in vivo. In contrast, introduction of the same antigen on dendritic cells matured with lipopolysaccharide (LPS) and anti-CD40 results in an IFN-γ-dominated response.
An increasing body of data, largely from in vitro studies, supports the notion that the type of innate receptors and the type of cell(s) being stimulated through them influence the cytokine milieu and the direction of the subsequent adaptive response. Thus, when stimulated with the fungal polysaccharide chitin, macrophages respond with TLR2-dependent IL-17 production.60 Some macrophage products, such as PGE2 and IL-1β, appear to synergistically promote a Th17 response.61 On the other hand, stimulation of macrophages with CpG oligodeoxynucleotides (CpGs) results in a TLR9-dependent production of IL-12, which, in turn, can drive a Th1 response.62 In contrast, CpG stimulation of CNS-infiltrating CD11c+ dendritic cells from EAE mice causes them to secrete IL-23 and to induce Th17 polarization.63 Although it was initially reported that, in the presence of LPS, dendritic cells produce IL-23 in a TLR4-dependent fashion, driving IL-17 production in T cells,64 it was subsequently demonstrated that LPS stimulation of TLR4 on its own does not drive Th17 production and that TGF-β signaling is required alongside TLR4 signaling for Th17 development.26 These findings underscore the complexity of innate differentiation factors and signals that independently or together affect effector T-cell differentiation. Dissection of these factors in vivo presents a daunting task. It is of note that in human patients with autoimmune disease the innate conditions surrounding initial exposure to antigen are unknown. However, if the concepts proposed above are correct, these events could affect the nature of the resulting disease and impact its treatment.
Just as Th1 and Th2 responses have long been thought to regulate one another, there is similar evidence regarding the relationship between Th1 and Th17. Lack of one of these cytokines in vivo appears to promote a response dominated by the other. This is particularly evident in each of the CFA-induced models of autoimmunity where IFN-γ-deficient mice develop an elevated Th17 response and exacerbated disease that features an increased neutrophilic infiltration at the site of inflammation.5,41,65 Conversely, IL-17 deficiency may permit emergence of Th1 as an effector response; in uveitic eyes of IL-17 knockout mice there are increased numbers of CD4+IFN-γ+ (Th1) cells, raising the possibility that these mice may use the Th1 response in lieu of the Th17 response to drive pathology.5 Recent data in the EAE model suggest that Th1 cells that produce ultra-high levels of IFN-γ may constitute a specialized population that suppresses Th17 development in the early stages of EAE (Nathan Karin, Technion, Israel Institute of Technology, Haifa, Israel, personal communication). The mechanisms of this reciprocal regulation are complex and include inhibition at the level of lineage-specific differentiation factors in that mediators of Th17 development may downregulate the Th1 response and vice versa. Thus, STAT3 activation has been shown to downregulate the expression of IL-12,66 whereas addition of IL-12 to cell cultures containing Th17-polarizing cytokines led to a dose-dependent reduction in the number of IL-17-producing cells.26 However, early IFN-γ production may regulate not only the Th1 but also the Th17 responses. Triggering of early IFN-γ production from NKT cells by the NKT-specific invariant T-cell receptor stimulant α-galactosyl ceramide (α-GalCer) cells results in subsequent attenuation of both Th1 and Th17 antigen-specific responses and inhibits induction of EAU.67
Although either Th1 or Th17 effector T cells can drive immune-mediated pathology, the disease induced by each effector population is distinct in terms of the type of inflammatory leukocytes recruited to the site of inflammation and in terms of the preferential tissue location of the pathology (that also affects clinical disease manifestations) (Table 1). Adoptive transfer of Th1-skewed cells resulted in a mononuclear infiltrate within the eyes of mice with EAU. In contrast, infusion of Th17-polarized cells resulted in a neutrophilic infiltrate as does genetic deficiency of IFN-γ, which results in an enhanced Th17 response.5,68 Similarly, Kroenke and colleagues demonstrated that adoptive transfer of Th1-polarized myelin-specific T cells results in heavy macrophage infiltration in EAE mice, whereas transfer of Th17-polarized cells leads to an infiltrate rich in neutrophils.69 In both models, the chemokine profile at the site of inflammation reflected the nature of the infiltrating cells, with monocyte chemoattractants CXCL9, 10, and 11 found in Th1-driven lesions and neutrophil-attracting chemokines CXCL1 and CXCL2 found in Th17-driven lesions.65,69 Similar observations were reported in CIA.70
Notably, the target tissue itself participates actively in creating the site-specific milieu by contributing to production of soluble mediators, including chemokines. Differences in cellular composition of the tissue could therefore be reflected in the local milieu, resulting in preferential attraction of Th1 versus Th17 and affecting the manifestations of disease. Stromnes and colleagues71 demonstrated that transfer of a Th17-predominant cell population resulted in an atypical form of EAE by which the cerebellum was inflamed and not the spinal cord (the normal inflammatory site of classical EAE).71 Symptomatically, this atypical EAE is characterized by dystonia of the limbs and tail. In contrast, adoptive transfer of a Th1-predominant population led to inflammation of the spinal cord and flaccid paralysis, hallmarks of classical EAE. In an independent but related study, Kroenke and colleagues69 found that inflammatory cells infiltrated different CNS sites depending on their Th1/Th17 polarization. Lees and colleagues72 demonstrated that these preferences in tissue localization are dependent on the tissue and are driven by the response of the tissue to IFN-γ produced by the infiltrating T cells.72
Both Th1 and Th17 cells co-localize within the region of inflammation5,54,73 and they may require each other for recruitment and/or entry to this region (Fig. 2). O’Connor and colleagues demonstrated that while both Th1 and Th17 cells were found in the CNS of mice with classical spinal cord-associated EAE, only myelin-specific IFN-γ-producing Th1 cells had the capacity to accumulate within the CNS and cause disease if transferred alone, whereas the Th17 cells could not.73 This study also suggested that in the course of EAE, Th1 cells accumulate first in the CNS and subsequently allow for the entry of pathogenic Th17 cells. In the lung, however, the reverse entry sequence has been reported in mice receiving a post-vaccination challenge of Mycobacterium tuberculosis; Th17 cells were recruited to the lung prior to the appearance of IFN-γ-producing memory cells and IL-17-induced chemokine expression was essential for the rapid accumulation of these Th1 effectors.74 This differential tissue preference is likely to be again a consequence of the target tissue response. It has been suggested that upon infection lung cells produce and secrete chemokines, such as CCL20, that specifically attract Th17 cells,75 whereas Th17 cells express CCR6, which is a receptor for CCL20.76 Conversely, astrocytes resident within the CNS express IP-10, a chemokine that attracts Th1 cells via the receptor CXCR3.77,78 These findings underscore the role of the tissue in affecting the site where inflammation occurs and provide evidence that Th1 and Th17 cells are not always functionally antagonistic but may in fact collaborate to accomplish some effector functions (Table 2).
While Th17 cells are considered to represent a distinct lineage, there is increasing evidence that the line between Th17 and Th1 may not be as clear as was initially thought. Not long after the first reports of the Th17 lineage, studies revealed subsets of cells producing both IFN-γ and IL-17 in the inflamed CNS79,80 as well as in the eye.5 Double producers may represent a transitional state between Th17 and Th1 cells. Using Th1-polarizing conditions and IL-17 reporter mice, Th17 cells have been shown by several investigators to be phenotypically unstable and to readily convert to a Th1 phenotype in vitro and in vivo. It appears, however, that under parallel conditions Th1 cells do not readily convert to Th17 cells.80–84 The molecular mechanisms involved in Th17/Th1 plasticity are complex and are only now starting to be elucidated.81,83,84 One of the mechanisms at play in this seemingly one-way street from Th17 to Th1 is that Th17 cells still express IL-12Rβ1 as part of IL-23R but Th1 cells do not express the unique IL-23R chain, which has expression maintained by IL-23 itself. When transferred to Th1-polarizing conditions and in the absence of IL-23, Th17 cells decrease synthesis of the IL-23R chain, removing the competition for the IL-12Rb2. This allows the IL-12Rb1 to associate with IL-12Rb2, reconstituting the heterodimeric IL-12 receptor and promoting synthesis of IFN-γ. In this process T cells either co-produce IFN-γ plus IL-17 or suppress IL-17 production and begin to produce IFN-γ (V.K. Kuchroo, Harvard Medical School, Boston, MA, personal communication).
In some cases a Th17 to Th1 conversion may play a functional role in pathogenesis. A recent diabetes study in NOD mice has suggested that, while adoptive transfer of polarized Th17 cells could promote pancreatic inflammation, their conversion to a Th1 phenotype was necessary for the induction of full-blown diabetes because only a blocking antibody to IFN-γ could block diabetes progression following adoptive transfer.85 This finding suggests that in addition to paving the way for sequential entry into the site of inflammation, a transition from the Th17 phenotype into an IFN-γ producing Th1-like cell can promote further disease. On the other hand, in the EAE model it has been suggested that conversion from a Th17 to a Th1 phenotype may have an important regulatory function. CNS-derived regulatory T cells (Tregs) were only able to suppress encephalitogenic effector T cells recovered just prior to EAE resolution.54 Because Th1 cells appear to be the dominant population just before the recovery process of EAE and appear to be more susceptible to Tregmediated suppression than Th17,79 such conversion of pathogenic Th17 cells into Th1 cells may represent a mechanism permitting resolution of disease. Plasticity of Th17 cells may also play a role in antitumor immunity. A recent study reported that adoptively transferred Th17 cells were superior to Th1 cells at inducing melanoma tumor regression. Unexpectedly, the enhanced antitumor effect was completely dependent on IFN-γ production by these cells. While not definitively excluding collaboration between the lineages as a factor, this finding may potentially implicate Th17 to Th1 conversion as an important step in the antitumor activity of Th17 effector cells.86,87
It has become clear that the relationship between the Th1 and Th17 lineages is much more intertwined and complex than was initially appreciated. While there is clear evidence that they counter-regulate each other, there is also increasing evidence of cooperation and even dependency between these two responses in effecting pathology. In view of the evidence that each response has the capacity to cause autoimmune disease independently of the other and that both may collaborate during an inflammatory response, efficient specific targeting of either Th1 or Th17 cells may prove potentially therapeutic for patients suffering from autoimmune diseases. However, because these responses may also regulate one another, it is conceivable that targeting one may cause an exacerbation of the other, leaving disease progression unaffected or even worsened. Alternatively, these studies may provide a case for the future development of therapies targeting both lineages. Better understanding of the ways in which Th1 and Th17 responses interact is essential for understanding disease pathogenesis and for determining appropriate therapeutic targets for autoimmunemediated inflammatory diseases.
Conflict of interest The authors declare no conflicts of interest.