The Role of IL-17 and Related Cytokines in Inflammatory Autoimmune Diseases

2.1. Helper T Cells

Various hematopoietic and lymphoid progenitors are mobilized from the bone marrow and initiate T cell development in the thymus. During this process, they express an antigen receptor (the T cell receptor, TCR), and most cells differentiate into CD4-positive T cells or CD8-positive T cells. After completion of the maturation process, CD8-positive T cells circulate throughout the body, acquiring cytotoxic functions. They contribute to immunological homeostasis by killing cells that have been infected by viruses as well as cancer cells. On the other hand, CD4-positive T cells are helper T cells. They exhibit an immunological regulatory function. Helper T cells have previously been divided into mainly two subsets (Figure 1) [8]. Th1 cells differentiate under the influence of IL-12 and mainly produce interferon-γ (IFN-γ). IFN-γ strongly activates macrophages, promoting the elimination of intracellular pathogens. In other words, it supports cellular immunity in the acquired immune system. On the other hand, Th2 cells differentiate under the influence of IL-4. Th2 cells support B cells through IL-4 production. As a result, the antibodies produced by B cells switch their class from IgM to IgG or IgE. By inducing the production of IgG or IgE, elimination of extracellular parasites (such as nematodes) is promoted. While cellular immunity is performed by Th1 cells, Th2 cells support humoral immunity. Thus, although derived from the same precursor cells, when activated by antigen stimuli, helper T cells differentiate into subsets with different properties due to the surrounding environmental factors (in particular, cytokines). The mechanism of differentiation has been previously analyzed in detail to clarify their mutually exclusive properties. Namely, IFN-γ suppresses the differentiation of Th2 cells, while IL-4 inhibits Th1 differentiation. Therefore, the functional imbalance between Th1 and Th2 is at the origin of various immunological diseases. For example, when there is a bias toward Th1, autoimmune diseases such as MS and RA are more likely to occur, while if Th2 is dominant then allergic reactions represented by pollinosis are provoked. However, energetic research in recent years identified subsets other than Th1 and Th2 cells [9]. Among these, the Th17 cell subset, which produces IL-17, contributes to autoimmune diseases accompanied by chronic immune and inflammatory reactions, working together with Th1 cells.

2.2. Interleukin-17

IL-17 is a cytokine whose gene was isolated from a rat-mouse T cell hybridoma in 1993. Since it displayed a high degree of homology with the HVS13 Herpes virus gene, it was thought to be a subtype of the cytotoxic T-lymphocyte-associated protein (CTLA) family of proteins and called CTLA-8 [10]. In 1995, it was recognized as a new cytokine and named IL-17, and, today, six homologous molecules are known (IL-17A through IL-17F). To date, most studies focused on IL-17A, IL-17E, and IL-17F. IL-17A and IL-17F are highly homologous and share receptors. Thus, they have extremely similar functions. IL-17E is also known as IL-25, and because it presents low homology with other molecules of the family and contributes to the induction of allergies, it is thought to have functions different from those of IL-17A [11]. This review focuses on IL-17A. Several types of immune cells produce IL-17A, and Th17 cells, the newly established subset of helper T cells, have received particular attention [12–14]. Through the analysis of the function of Th17 cells in autoimmune diseases, there has been significant progress in the understanding of the biological significance of IL-17.

2.3. IL-17 Receptors and Their Signaling

IL-17 receptor A (IL-17RA) was identified as a new cytokine receptor for IL-17A and was found to be a member of the cytokine receptor family [15]. The IL-17 receptor family now consists of five members (IL-17RA, IL-17RB, IL-17RC, IL-17RD, and IL-17RE), all of which share sequence homology. All these receptors contain a fibronectin III-like domain in their extracellular region and a SEF/IL-17R (SEFIR) domain in their intracellular region [16]. IL-17RA had been long considered as the receptor of IL-17. However, newly emerged evidence suggests that IL-17RA is likely a common receptor used by the IL-17 family cytokines. Other receptors including IL-17RB, IL-17RC, and IL-17RE have been identified as specific receptors for IL-17E, IL-17A, and IL-17F and IL-17C, respectively. IL-17RD, originally identified as a negative regulator in fibroblast growth factor signaling, has recently been found to regulate IL-17A signaling [17, 18]. Recent studies have shown that IL-17A and IL-17F, believed to be predominantly expressed in Th17 cells, signal through a heteromeric receptor complex. This complex consists of IL-17RA and IL-17RC, which are single transmembrane proteins and ubiquitously expressed in various cell types, including epithelial cells, fibroblasts, and astrocytes [19–22]. It has been well documented that IL-17A and IL-17F can induce proinflammatory gene expression both alone and in synergy with TNFα, IL-6, G-CSF, IL-1, CXCL1, CCL20, and matrix metalloproteases [23–25]. IL-17 upregulates these proinflammatory genes through the activation of NF-κB, MAPK, and C/EBP cascades [26]. Other studies demonstrated that IL-17 also activates the Jak-Stat and Jak-PI3 K pathways [27, 28]. In addition to altering gene expression, IL-17 stimulation can stabilize mRNAs. Normally, mRNA splicing factor (SF) is bound to mRNA to mediate its degradation. Upon IL-17 stimulation, mRNA is dissociated and stabilized [29, 30].

Adaptor protein Act1 and tumor necrosis factor receptor-associated factor (TRAF) regulate these molecular mechanisms after activation of IL-17R. Act1 was originally discovered as an NF-κB activator [31, 32]. Several studies demonstrated that Act1 is recruited to the IL-17 receptor complex through the homotypic interactions of the SEFIR domains upon IL-17 stimulation [33, 34]. Act1 deficiency results in loss of IL-17-dependent NF-κB activation and proinflammatory cytokine production. Following Act1 binding to the receptor complex, TRAF6 is recruited through interaction with the Act1 TRAF binding motif [35]. TRAF6 further activates downstream TRAF6-dependent TAK1 for NF-κB activation.

IL-17-mediated mRNA stability of CXCL1 requires Act1 but not TRAF6, suggesting that Act1 mediates both TRAF6-dependent and TRAF6-independent pathways in IL-17R signaling [29]. In the TRAF6-independent pathway, TRAF2 and TRAF5 are required for IL-17-induced mRNA stabilization of CXCL1 [30]. Act1-TRAF5-TRAF2-SF complex formation induced by IL-17 prevents SF binding to CXCL1 mRNA for degradation. Phosphorylation of Act1 at Ser-311 was shown to be required for its association with TRAF2-TRAF5 complex. By mass spectrometry analysis, Act1 was found to be phosphorylated directly by the kinase IKKε at Ser-311 upon exposure to IL-17. Consistently, IKKε deficiency also prevented the formation of Act1-TRAF2-TRAF5 complex without disrupting interaction between TRAF6 and Act1, suggesting that phosphorylation of Act1 is necessary for the formation of the mRNA stabilization cascade. Thus, Act1 serves as a receptor proximal anchor platform for the initiation of two independent pathways activated by IL-17: (1) the TRAF6-dependent cascade for signaling and (2) TRAF2-TRAF5-dependent cascade for stabilization of mRNA.

Regulation of IL-17R pathways is important in the control of IL-17-mediated inflammation. As in intracellular mechanisms, fundamental features of inflammation have been revealed using mice lacking IL-17R subunits. IL-17RA-deficient fibroblasts failed to respond to IL-17A or IL-17F stimulation [36]. Subsequent studies showed that IL-17RC interacts with IL-17RA and it can bind to IL-17A or IL-17F. IL-17RC-deficient mice also failed to induce downstream gene expression in response to IL-17A and IL-17F and develop much milder disease than wild-type mice in experimental autoimmune encephalomyelitis [37–39]. These results suggest that the IL-17 receptor complex has a significant role in biological and pathological functions [39] and is a candidate therapeutic target. Therefore, identification of new compounds to block the activation of IL-17R is a powerful approach to prevent IL-17-mediated pathology. A monoclonal antibody targeting IL-17R would be a potential treatment to investigate this therapeutic tool for autoimmune diseases. Brodalumab, previously known as AMG827, is a human monoclonal antibody that binds to the human IL-17RA and blocks the biological activities of IL-17A and IL-17F. This antibody is a broad spectrum inhibitor of IL-17-mediated signaling pathways compared to other IL-17-targeted therapy. Brodalumab is currently being investigated in different phases of clinical trials for psoriasis (phase II) and for RA (phase I/II) [40, 41]. In a phase II study, brodalumab showed significant improvement in severe psoriasis compared to placebo [40]. In these preclinical studies, IL-17R targeted therapy did not show any major safety concern. However, further trials are warranted to confirm the safety profile of this therapy. IL-17 expresses protective properties against extracellular pathogens [42, 43]. Future trials with a large number of patients will provide more insight into the safety profile. Other therapeutic strategies against IL-17 are discussed below.

3.1. Th17 Cells in EAE

MS is an autoimmune disease accompanied by chronic neuroinflammation that causes demyelination in the CNS. Accumulation of helper T cells is observed in the cerebrospinal fluid of patients. The EAE mouse model that recapitulates MS is an experimental system in which pathogenic helper T cells can be induced by administering antigens in the CNS tissue, resulting in limb paralysis. The pathophysiological analysis of EAE is useful for understanding MS. MS-related genes have been proposed by establishing and screening the disease-related genome analysis (genome-wide association studies, GWAS) database [44]. The results suggest that specific major histocompatibility complex (MHC) class II genes are associated with a significantly high risk for the onset of MS. Moreover, although somewhat less important than the MHC, a group of genes related to differentiation and activation of helper T cells also showed a significant relationship with MS. Thus, it is believed that this disease of the CNS is a T cell-mediated disease.

3.2. Th17 in RA and Psoriasis

As for MS and EAE, Th17 cells strongly contribute to RA. Since Th17 cells cannot be separated from RA and MS disease conditions, the role and function of IL-17 have been analyzed in detail. The important clinical issue for RA is bone and joint destruction. Since bone destruction is directly related to changes in the joint structure, treatment for prevention of joint destruction is very important. Osteoclasts are the only cells that break down the bone, and they are a specially differentiated type of macrophage. Osteoclasts differentiate from monocytes, and receptor activator of nuclear factor kappa-B ligand (RANKL) promotes this process [83, 84]. Studies focused on how helper T cells that infiltrate the joints induce osteoclastogenesis from monocytes. Helper T cells that produce IL-17 have been found in the synovial tissue of patients with RA [85]. IL-17 stimulates the osteoblasts in the joints, inducing RANKL expression. Then, through the interaction with osteoblasts, monocytes respond to RANKL and mature into osteoclasts. After this mechanism was reported, interest in IL-17 for RA and its animal models grew [86, 87]. The identity of the helper T cells that regulate IL-17 production in the synovium remained unclear, but, after the discovery of Th17 cells as a new subset in 2006, the RANKL-dependent induction mechanism of osteoclasts via Th17 cells in RA became clear [88]. In RA and CIA, continuing inflammation is an important disease condition, similar to bone destruction. IL-22 produced by Th17 cells certainly regulates chronic inflammation. IL-22 stimulates synovial fibroblasts to induce cell proliferation and the production of inflammatory chemokines [89]. Studies using a mouse model also showed that IL-22 induces osteoclastogenesis [90]. Moreover, IL-6 produced by fibroblasts responding to IL-17 derived from Th17 cells amplifies inflammation [79]. IL-6 stimulates synovial tissue in an autocrine manner, worsening the condition by amplification cycle that releases inflammatory mediators. Thus, IL-17 maintains the inflammatory cycle via downstream cytokines. It has recently been reported that IL-17 in the synovial tissue is not derived from Th17 cells, but rather from mast cells [91]. That does not change the importance of IL-17 for the disease condition, but the possibility remains that the role of Th17 cells in this disease will be revised.

Psoriasis is a chronic inflammatory skin disease characterized by hyperproliferation and abnormal differentiation of epidermal cells. Pronounced acanthosis and inflammatory infiltration such as of neutrophils and lymphocytes are observed in lesions. As cyclosporin, a calcineurin inhibitor, shows therapeutic effectiveness, it is thought that T cells contribute to psoriasis pathology. Recent studies clarified that Th17 cells contribute to the onset of psoriasis [92, 93]. Th17 cells that have infiltrated the skin produce not just IL-17, but also IL-22. IL-22 stimulates keratocytes in the skin, leading to the activation of STAT3, followed by the proliferation of keratinocytes and acanthosis [94]. IL-17 and IL-22 induce keratinocytes to express CXCL1 and CXCL8, inducing further cell infiltration [94–98]. Thus, a positive psoriasis feedback loop is formed. A contribution of IL-36 to the detailed mechanism of this positive feedback loop has been suggested [99]. IL-36 is produced by keratinocytes and stimulates resident DCs that are normally present in the skin. In response to IL-36, DCs express IL-23, which appears to increase the pathogenicity of Th17 cells [100]. In the normal skin, an IL-36R antagonist is expressed, to compete with IL-36 stimulation [101]. However, it is thought that, as the disease condition progresses, the suppression by the IL-36R antagonist is abrogated.

3.3. Plasticity of Pathogenic Helper T Cells

In chronic inflammation, IL-23 stimulation confers higher pathogenicity on Th17 cells, and, in this process, not just IL-17, but also IFNγ, IL-22, and GM-CSF are produced. In a recent study, based on the way functional cytokines are produced, helper T cells have been divided into multiple subsets such as Th9 cells and Th22 cells [2]. Th1 cells and Th2 cells stably maintain their properties. In contrast, it seems that Th17 cells infiltrated into CNS lesions produce various cytokines and differentiate from Th17 cells into other subsets [102]. Studies using an IL-17 reporter mouse indicated that, during EAE onset, cells that produced IL-17 were converted into cells that produced IFNγ [103]. Based on this result, a model was proposed in which Th17 cells redifferentiated either into Th17/Th1 hybrid cells, or into a type of Th1 cells that stopped producing IL-17 (ex-Th17). This model may explain the presence of Th1/Th17 hybrid cells in the CNS from patients with MS. Moreover, these cells producing IFNγ, which may be ex-Th17 cells, are more pathogenic than Th17 cells. IFNγ can be produced by conventional Th1 cells and ex-Th17 cells. However, conventional Th1 cells are not exposed to IL-23 and, therefore, do not express IL-1R, while ex-Th17 cells exposed to IL-23 in the past can express IL-1R. Therefore, the expression of IL-1R helps distinguishing between IFNγ-producing ex-Th17 cells and conventional Th1 cells. Since GM-CSF is encephalitogenic, a new subset of cells has been named ThGM-CSF [56, 57]. According to this model, it is highly possible that GM-CSF-producing helper T cells acquire the ability to produce GM-CSF after Th17 cells stop producing IL-17. In fact, helper T cells that produce both IL-17 and GM-CSF have been observed [104]. Additionally, there are cases where other subsets turn into Th17 cells. TGFβ, which is necessary for the differentiation of Th17 cells, is a cytokine that induces the differentiation of Treg cells. Komatsu et al. using a CIA mouse model showed that Foxp3-positive Treg cells in the joint synovia redifferentiated into Th17 cells (IL-17-producing ex-Treg cells) [105]. IL-17-producing ex-Treg cells express RANKL and cause strong osteoclast differentiation compared to the normal type of Th17 cells. These monocytes interact directly with IL-17-producing ex-Treg cells, not via synovial fibroblasts.