IL-6–gp130–STAT3 in T cells directs the development of IL-17+ Th with a minimum effect on that of Treg in the steady state (original) (raw)

Journal Article

,

1Laboratory of Developmental Immunology, Graduate School of Frontier Biosciences and Graduate School of Medicine, Osaka University, Osaka, Japan

Search for other works by this author on:

,

1Laboratory of Developmental Immunology, Graduate School of Frontier Biosciences and Graduate School of Medicine, Osaka University, Osaka, Japan

Search for other works by this author on:

,

1Laboratory of Developmental Immunology, Graduate School of Frontier Biosciences and Graduate School of Medicine, Osaka University, Osaka, Japan

Search for other works by this author on:

,

1Laboratory of Developmental Immunology, Graduate School of Frontier Biosciences and Graduate School of Medicine, Osaka University, Osaka, Japan

Search for other works by this author on:

,

1Laboratory of Developmental Immunology, Graduate School of Frontier Biosciences and Graduate School of Medicine, Osaka University, Osaka, Japan

Search for other works by this author on:

,

2Santen Pharmaceutical Co., Osaka, Japan

Search for other works by this author on:

,

2Santen Pharmaceutical Co., Osaka, Japan

Search for other works by this author on:

,

1Laboratory of Developmental Immunology, Graduate School of Frontier Biosciences and Graduate School of Medicine, Osaka University, Osaka, Japan

Search for other works by this author on:

,

3Department of Pathology, University of Massachusetts Medical School, Worcester, MA, USA

Search for other works by this author on:

,

4Preclinical Research and Development, Merck KGaA, 64293 Darmstadt, Germany

Search for other works by this author on:

... Show more

• PDF
• Split View
• • Article contents
  • Figures & tables
  • Video
  • Audio
  • Supplementary Data
• Cite

Cite

Mika Nishihara, Hideki Ogura, Naoko Ueda, Mineko Tsuruoka, Chika Kitabayashi, Fumio Tsuji, Hiroyuki Aono, Katsuhiko Ishihara, Eric Huseby, Ulrich A. K. Betz, Masaaki Murakami, Toshio Hirano, IL-6–gp130–STAT3 in T cells directs the development of IL-17+ Th with a minimum effect on that of Treg in the steady state, International Immunology, Volume 19, Issue 6, June 2007, Pages 695–702, https://doi.org/10.1093/intimm/dxm045
Close

Navbar Search Filter Mobile Enter search term Search

Abstract

IL-17-producing Th (Th17) comprise a distinct lineage of pro-inflammatory Th that are major contributors to autoimmune diseases. Treatment with IL-6 and transforming growth factor β (TGFβ) induces naive CD4+ T cells to generate Th17, which also requires expression of the IL-6/TGFβ target RORγt. We reported that IL-6 transduces two signaling pathways via tyrosine redidues of the signal transducer gp130: one depends on signal transducers and activators of transcription (STAT)-3 activation and the other on Src homology region 2 domain-containing phosphatase 2 (SHP2)/Grb2 associated binder (Gab)/mitogen-activated protein kinase (MAPK) activation. Here, we showed that CD4+ T cells carrying a mutant gp130 that transduces the SHP2/Gab/MAPK pathway but not the STAT3-mediated one failed to develop into Th17, while CD4+ T cells whose mutant gp130 transduces the STAT3 signal only generated Th17, indicating that IL-6 acts directly on T cells through the tyrosine residues of gp130 required for STAT3 activation to promote the development of Th17. Moreover, we found that gp130–STAT3 pathway is essential for Th17 development and for the expression of RORγt by using T cells specifically lacking gp130 and STAT3. Noteworthy is that the regulatory T cell (Treg) percentages and numbers were comparable between all mutant mice we tested in vivo, although we showed that IL-6–gp130–STAT3 pathway suppressed Treg development in vitro. Thus, we conclude that IL-6 acts directly to promote the development of Th17 by activating the T cell gp130–STAT3 pathway but has a minimum effect on Treg development at least in the steady state in vivo. Therefore, blockade of IL-6–gp130–STAT3 pathway in CD4+ T cells could be a good target for controlling unwanted Th17-mediated immune responses including autoimmune diseases.

Introduction

The Janus kinase (Jak)–signal transducers and activators of transcription (STAT) pathway was originally discovered through the study of IFN-induced signal transduction. Since then, a large number of cytokines, hormones and growth factors have been found to activate Jaks and STATs. Upon ligand binding, the STAT family is activated to form dimers, which translocate to the nucleus and bind to specific response elements in the promoters of target genes.

Until the discovery of the recently described Th17 lineage, the adaptive CD4+ T-cell responses have been encompassed by the Th1–Th2 paradigm (1, 2). The development of Th1, which enhance the clearance of certain intracellular pathogens, is dependent on IFNγ and IL-12 (3, 4). It is now known that Th1 generation is mediated by IFNγR signaling via STAT1, driving Tbet expression (5, 6). The development of Th2, which enhance the clearance of parasites or the generation of Igs, is dependent on IL-4 (7, 8). The Th1 and Th2 populations can also be categorized by their distinct requirements for STAT molecules: STAT4 and STAT1 for Th1 (5, 6, 9, 10) and STAT6 for Th2 (11, 12). Mice deficient in IL-17 are resistant to collagen-induced arthritis (CIA) (13). The development of the IL-17-producing T cells, now termed Th17, was initially shown to depend on the presence, during antigen stimulation, of IL-23 produced by the antigen-presenting cells (14–17). Although IL-23 has a key role in Th17-mediated inflammation in vivo (18), recent studies have demonstrated that the in vitro polarization of naive CD4+ T cells toward the Th17 lineage requires a combination of T cell antigen receptor stimulation and the cytokines transforming growth factor β (TGFβ) and IL-6, but that it is independent of IL-23 (19, 20). Additionally, it was previously reported that in vivo polarization of naive T cells toward the Th17 lineage requires IL-6 and TGFβ (19, 21). Importantly, it is reported that IL-6 signaling suppressed in vitro regulatory T cell (Treg) development, suggesting reciprocal development pathways for the generation of Th17 and Tregs (19). However, it has not been shown whether these reciprocal development pathways also exist in in vivo situation. Recently, RORγt was identified as a critical target of TGFβ and IL-6 for the development of Th17 (22). However, it is not known if any STAT family ‘in T cells’ is critically involved in Th17 development, although STAT3 is the major STAT that is activated by IL-6 and STAT3 is previously known to bind both the IL-17A and IL-17F promoters (18) and STAT3 depletion in hematopoietic stem cells as well as endothelial cells using Tie2-Cre mice (23) decreased Th17 development (24).

The IL-6R consists of a ligand-binding α-chain and the signal transducer gp130, which is shared among the receptors for the other IL-6 family cytokines (25, 26). Binding of the IL-6 family cytokines to their receptors activates Jak, leading to the recruitment of two important signal-transducing molecules: Src homology region 2 domain-containing phosphatase 2 (SHP2) (protein tyrosine phosphatase 2) and STAT3. The gp130 subunit contains six tyrosines in its cytoplasmic region and transduces two major signaling pathways, which are dependent on these tyrosine residues: (i) Tyr759 for human gp130 (Tyr754 for mouse) is required for the tyrosine phosphorylation of SHP2 and (ii) any one of four tyrosines in the C-terminus (Tyr767, Tyr814, Tyr905 and Tyr915), which have a glutamine at position 3 of the tyrosine motif (YXXQ), is required for the tyrosine phosphorylation of STAT3 (25, 26). Tyr759 is required for the tyrosine phosphorylation of SHP2 and the activation of Gab, extracellular signal-regulated kinase (ERK) and mitogen-activated protein kinase (MAPK). It is also shown to have a negative role in STAT3-mediated biological actions mediated by a STAT3 inhibitor, suppressor of cytokine signaling 3 (SOCS3) or SHP2 itself (25). We previously established a series of knock-in mice expressing mutant gp130 defective in the SHP2/Gab/MAPK pathway (the F759 mouse), the STAT3-dependent pathway (the FxxQ mouse) or both (27). We observed that the F759 mouse, lacking the SHP2/Gab/MAPK pathway, has splenomegaly and lymphadenopathy from an early age and develops autoimmune arthritis in later life (28, 29). However, which IL-6-mediated pathway is critical for the development of Th17, i.e. the SHP2/Gab/MAPK or the STAT3-dependent pathway in T cells, has not been studied. We show here that the IL-6–gp130-mediated STAT3 pathway in T cells is required for RORγt expression and the development of Th17 with a minimum effect on in vivo Treg development at least in the steady state.

Methods

Mice

C57BL/6 mice were purchased from Japan SLC (Shizuoka, Japan). gp130flox/flox mice were established previously (30) and crossed with lckCre transgenic mice. DBA-backcrossed F759 mice (more than eight times) were established by us. C57BL/6-backcrossed lckCre transgenic mice and STAT3flox/flox mice were obtained from S. Akira (31) (Osaka University). The gp130F759/F759 knock-in mice, which carry a human version of gp130 (S710L), were established previously (27, 32). IL-6 knockout (KO) mice were obtained from M. Kopf (Max-Planck-Institut fur Immunobiologie) via Y. Iwakura (University of Tokyo) (33, 34). Mice were maintained in the animal facilities of the Medical Sciences, Osaka University, under specific pathogen-free conditions. Animal experiments were performed following the guidelines of the Institutional Animal Care and Use Committees of the Graduate School of Frontier Biosciences and the Graduate School of Medicine of Osaka University. We just used 7- to 12-week old young animals for all the experiments without description in the figure legends.

Fetal liver transplantation

To analyze the T cells in gp130FxxQ/FxxQ mice, which die after birth in neonate state, we transplanted fetal liver cells from gp130FxxQ/FxxQ embryos at 14.5 days into lethally irradiated (950 rad) C57BL/6 SJL mice, as described previously (32).

Antibodies and reagents

The following mAbs were used: allophycocyanin (APC)-conjugated anti-CD4 (BioLegend), anti-CD8 (eBiosciences), anti-CD19 (eBiosciences), IgG1 (BD), anti-IL-4 (BD) and anti-IFNγ (BD); FITC-conjugated anti-CD4, anti-CD8, anti-B220, anti-Foxp3 (eBiosciences) and IgG2a (eBiosciences); PE-conjugated anti-CD4, anti-CD8, anti-CD44 (eBiosciences), anti-CD25 (eBiosciences), anti-IL-17 (eBiosciences) and IgG1 (eBiosciences); Cy5-conjugated anti-CD44 (eBiosciences), anti-CD8 (eBiosciences), streptavidin and Pacific Blue conjugate (Invitrogen) and biotin-conjugated anti-CD8, anti-CD19, anti-CD11c, anti-B220, anti-NK1.1 and anti-CD45.1 (eBiosciences).

FACS analysis

Cultured or fresh whole or CD4+ T cells from spleen and lymph nodes were prepared and incubated with fluorescence-conjugated antibodies (for specific staining and dump staining). FACS was performed on a FACSCalibur (Becton Dickinson) or a CyAn flow cytometer (Dako Cytomation).

Cell preparation and cell sorting

The lymph nodes and spleen from wild-type control or mutant mice were harvested and naive CD4+CD44Low T cells were purified using a MoFlo cell sorter (Dako Cytomation). The purity of the T cells was consistently >99%.

In vitro T-cell differentiation

Naive, sorted CD4+ T cells (5 × 105 ml−1) were cultured in vitro with or without anti-CD3 antibody (1 μg ml−1) (eBiosciences), IL-6 (1 μg ml−1) or TGFβ (25 ng ml−1) in the presence of bone marrow-derived dendritic cells (1 × 105 ml−1) for 5 days.

Intracellular staining

In vitro assay.

The recommended amount of Golgiplug (BD) was added to the culture during the last 6 h. Cells were stained with APC-conjugated anti-CD4 antibody, biotin-conjugated anti-CD19, anti-NK1.1 and anti-CD45.1 antibodies (eBiosciences), followed by streptavidin–Pacific Blue treatment (Invitrogen). Next, the cells were fixed and permeabilized according to the manufacturer’s directions (eBiosciences) and then stained with FITC-conjugated anti-Foxp3 and PE-conjugated anti-IL-17 (TC11-18H10) antibodies (eBiosciences). Data were acquired on a CyAn flow cytometer (Dako Cytomation) and analyzed with FlowJo software.

In vivo assay.

For Foxp3 staining, T cells from the lymph nodes and spleen were stained with APC-conjugated anti-CD4 (eBiosciences) antibody. The cells were fixed and permeabilized as above and then stained with FITC-conjugated anti-Foxp3 (eBiosciences) and biotin-conjugated anti-CD19 and anti-NK1.1 antibodies (eBiosciences) followed by streptavidin–Pacific Blue treatment (Invitrogen). For IL-17 staining, T cells from the lymph nodes and spleen were stimulated with plate-coated anti-CD3 antibody (20 μg ml−1) (eBiosciences) and anti-CD28 antibody (10 μg ml−1) (eBiosciences) for 6 h. The recommended amount of Golgiplug (BD) was added to the culture from the beginning of the incubation. The resulted cells were stained with FITC-conjugated anti-CD44 (eBiosciences), APC-conjugated anti-CD4 antibody and biotin-conjugated anti-CD19, anti-NK1.1 and anti-CD45.1 antibodies (eBiosciences) followed by streptavidin–Pacific Blue treatment (Invitrogen). The cells were then fixed in 4% PFA, permeabilized with PBS supplemented with 0.2% saponin and 0.05% sodium azide and stained with PE-conjugated anti-IL-17 (TC11-18H10) antibody (eBiosciences). Data were acquired on a CyAn (Dako Cytomation) and analyzed with FlowJo (for Foxp3) and Summit (for IL-17) software.

Real-time PCR

Total RNA was extracted from cells using an RNA isolation kit (Sigma). Complementary DNA was prepared as recommended and used as the template for quantitative PCR. The expression of RORγt was evaluated using specific primers for RORγt (5′-GGAGCTCTGCCAGAATGACC-3′ and 5′-CAAGGCTCGAAACAGCTCCAC-3′) on a GeneAmp 5500 Sequence Detection System (Applied Biosystems). Expression was normalized to the expression of the housekeeping gene HPRT, as described previously (35).

Statistical testing

Student’s _t_-test (two tailed) was used for statistical testing between two groups.

CIA

CIA was brought about in DBA/1 J mice (Japan SLC) essentially according to the method previously described (36). Mice were injected intra-dermally into the base of the tail with 200 μg of bovine type II collagen (CII) emulsified in CFA. Three weeks after the initial injection, a booster injection of 200 μg of bovine CII emulsified in CFA was performed intra-dermally into the base of the tail. Evaluation of clinical arthritis activity was carried out every 3 days from the second immunization for 27 days and its severity in the metacarpophalangeal wrist, metatarsophalangeal and ankle joints was scored as 0 = no arthritis, 1 = small degree of arthritis, 2 = light swelling, 3 = medium swelling and 4 = severe swelling and non-weight bearing. The arthritic score was the sum of the scores of all joints involved.

Measurement of IL-17 concentration in serum

IL-17 concentrations in sera were measured by ELISA kit (eBioscience) or Bio-plex system (Bio-Rad) according to the manufacture’s protocols.

Results and discussions

STAT3-binding tyrosine residues but not the tyrosine for SHP2/Gab/ERK/MAPK activation in gp130 play a role in Th17 development both in vitro and in vivo

To investigate which IL-6–gp130-mediated signaling pathway is critical for Th17 development, we sorted naive CD4+ T cells from two types of knock-in mice: FxxQ mice, which transduce the SHP2/Gab/ERK/MAPK pathway but not the STAT3-mediated one via their mutant gp130, and F759 mice, which have a mutant gp130 that transduces the STAT3-dependent pathway but not the SHP2/Gab/ERK/MAPK one (25–27).

We sorted naive CD4+ T cells from these mice and cultured them to generate Th17. FxxQ CD4+ T cells expressed almost no IL-17 molecules, while F759 T cells expressed significantly more IL-17 than did CD4+ T cells from control animals in the culture (Fig. 1a and b). The percentage of Foxp3+CD4+ T cells increased in the culture of FxxQ CD4+ T cells with IL-6 in the presence of TGFβ (Fig. 1a and c). Since IL-6–gp130-mediated STAT3 activation is enhanced in F759 cells (data not shown) (27, 32) and defective in FxxQ CD4+ T cells, these results strongly suggested a critical role for STAT3 in the development of Th17 as well as reciprocal development pathways for the development of Th17 and Tregs in vitro.

Fig. 1.

gp130-mediated STAT3 signaling but not SHP2/Gab/ERK/MAPK one in T cells is important for Th17 development in vitro. (a–c) MoFlo-sorted naive CD4+CD44Low T cells from FxxQ, F759, gp130flox/flox–lckCre or STAT3flox/flox–lckCre mice were stimulated with soluble anti-CD3, bone marrow-derived dendritic cell and the indicated cytokines. Five days after the activation, the cells were subjected to intracellular staining. (a) The dot plots represent IL-17 or Foxp3 expression in CD4+ cells. (b) The percentage of IL-17-producing CD4+ T cells was less in FxxQ, gp130flox/flox–lckCre or STAT3flox/flox–lckCre CD4+ T cells than in control CD4 T cells in the presence of IL-6 plus TGFβ (IL-17: P = 0.0026, 0.00000014 or 0.00012, respectively), while the percentage of IL-17-producing CD4+ T cells was greater in F759 CD4+ T cells in the presence of IL-6 plus TGFβ than in control CD4+ T cells (IL-17: P = 0.014). (c) The percentage of Foxp3+CD4+ T cells was greater in the presence of IL-6 plus TGFβ than in CD4+ T cells from FxxQ, gp130flox/flox–lckCre or STAT3flox/flox–lckCre mice (Foxp3: P = 0.029, 0.013 or 0.00014, respectively). (d) RORγt mRNA expression following stimulation with or without IL-6 in the presence or absence of TGFβ for 96 h. The relative expression levels were measured by quantitative real-time reverse transcription–PCR and normalized to the HPRT expression level, using the standard curve method. RORγt mRNA expression decreased in CD4+ T cells from FxxQ, gp130flox/flox–lckCre or STAT3flox/flox–lckCre mice compared with controls (RORγt: P = 0.001, 0.00076 or 0.027).

We confirmed that a critical role of IL-6 for the development of Th17 in vivo (Fig. 2a and b), although we observed that a comparable level of Tregs (Foxp3+CD4+ T cells) in IL-6KO comparing to wild-type control mice (Fig. 2c–e). Next, we asked whether the same signaling pathway, via gp130, is required for the in vivo development of Th17. We approached this question by examining Th17 development in the FxxQ mice, and we observed almost no Th17 in their lymph nodes and/or spleen (Fig. 2a and b). The percentage and number of Foxp3+CD4+ T cells in the FxxQ mice were comparable to those of wild-type controls (Fig. 2c–e). The F759 mice showed a comparable number of and a reduced percentage of Th17 compared with wild type (Fig. 2a and b). Again, the percentage and number of Foxp3+CD4+ T cells in the F759 mice were comparable to those of wild-type controls (Fig. 2c–e). Since the number of memory/activated CD44highCD4+ as well as total CD4+ T cells is increased in the F759 mice (data not shown), but the absolute number of Th17 (all of them are CD44highCD4+) remains the same as wild-type mice, this suggests an increase in other Th populations more in F759 mice comparing to Th17. Therefore, we analyzed Th1 and Th2 populations in F759 mice and showed that both percentage and the number of Th1 population judged by the expression of IFNγ but not Th2 population were increased (Supplementary Figure 1, available at International Immunology Online and data not shown). Taken together, all the in vitro and in vivo data demonstrated that STAT3-binding tyrosine residues in gp130, but not the tyrosine for SHP2/Gab/ERK/MAPK activation, are important in Th17 development. Additionally, we suggested that IL-6–gp130–STAT3 pathway had a minimum effect on the development of Treg in the steady state in vivo, although it suppressed the development in vitro.

Fig. 2.

gp130-mediated STAT3 signaling but not SHP2/Gab/ERK/MAPK one in T cells is important for Th17 development in vivo. (a and b) T cells from the spleen and lymph nodes of IL-6KO, FxxQ, F759, gp130flox/flox–lckCre or STAT3flox/flox–lckCre mice and wild-type controls were harvested and stimulated with anti-CD3 and anti-CD28 for 6 h. The cells were then subjected to intracellular staining. (a) The percentage of IL-17-producing CD4+ T cells decreased in the IL-6KO, FxxQ, F759, gp130flox/flox–lckCre or STAT3flox/flox–lckCre mice compared with controls (IL-17: P = 0.0000041, 0.0000047, 0.021, 0.015 or 0.049). (b) The number of IL-17-producing CD4+ T cells decreased in the IL-6KO, FxxQ, gp130flox/flox–lckCre or STAT3flox/flox–lckCre mice compared with controls (IL-17: P = 0.00035, 0.0019, 0.011 or 0.0085). (c–e) T cells from the spleen and lymph nodes of IL-6KO, FxxQ, F759, gp130flox/flox–lckCre or STAT3flox/flox–lckCre mice and wild-type controls were harvested and stimulated with anti-CD3 for 6 h. The cells were then subjected to intracellular staining. (c) The histograms represent Foxp3 expression in CD4+ cells. The dotted lines represent control Ig staining.

gp130 and STAT3 in T cells are essential for Th17 development

All the results described above strongly suggested that the direct action of IL-6 on T cells, accompanied by STAT3 activation, is required for Th17 development. To prove this, we used the progeny of gp130flox/flox (30) and STAT3flox/flox mice (31) crossed with T cell-specific lckCre mice. Yang et al. (24) recently showed that STAT3 depletion in endothelial cells and hematopoietic stem cells using Tie2-Cre mice (23) showed the decreased level of Th17, but this study cannot exclude a possibility that STAT3 in non-T cell population including endothelial cells indirectly plays a role for generation of Th17 population especially in vivo. Therefore, we examined a role of STAT3 molecules in T cells using lck-Cre mice for Th17 generation.

We first investigated the in vitro differentiation of Th17 in the presence and absence of IL-6 and TGFβ using naive CD4+ T cells from gp130flox/flox– and STAT3flox/flox–lckCre mice. Almost no Th17 developed from the gp130- and STAT3-deficient CD4+ T cells and the percentage of Foxp3+ cells increased in vitro, even in the presence of IL-6 in these CD4+ T cells (Fig. 1a–c).

Recently, RORγt was reported to be a critical target of IL-6 and TGFβ signaling to generate Th17 (22). Stimulation with IL-6 plus TGFβ in the presence of T cell antigen receptor-mediated signaling synergistically increases RORγt expression in naive CD4+ T cells (22). Moreover, RORγt expression is dramatically lower in the absence of either of these cytokines (22). The expression of RORγt was diminished in the STAT3-deficient CD4+ T cells (Fig. 1d) as well as FxxQ and gp130-deficient CD4+ T cells (Fig. 1d), while RORγt induction in F759 CD4+ T cells enhanced compared with that in wild-type controls (Fig. 1d). Because it was reported that STAT3 binds both the IL-17A and IL-17F promoters (18), STAT3 might also play a role for the induction of IL-17 mRNA. Together with the results presented here, the existing evidence supports the idea that the STAT3-dependent expression of RORγt provides at least in part the molecular basis for the critical role played by IL-6–gp130–STAT3 signaling in promoting the development of Th17 from CD4+ T cells.

Importantly, we showed that the in vivo development of Th17 significantly decreased in gp130flox/flox– and STAT3flox/flox–lckCre mice, (Fig. 2a and b) while we showed that a comparable level of Tregs in these mutants comparing to wild-type control mice (Fig. 2c–e). Taken together, all these results establish a critical role for the IL-6–gp130–STAT3 pathway in vivo in the development of CD4+ T cells into Th17 in vitro and in vivo.

Furthermore, our results showed that IL-6–gp130–STAT3 pathway in T cells had a minimum effect on in vivo Treg development. It is reported that Treg development takes place primarily in the thymus (37, 38), but peripheral development, i.e. reciprocal generation of Treg versus Th17 from mature peripheral CD4+ T cells is frequently suggested based entirely on in vitro data. The results here demonstrated that the reciprocal developmental pathways for the generation of Th17 and Tregs observed in vitro do not operate in vivo at least in the steady state. Moreover, these data suggests that the major development of Treg occurs in the thymus via IL-6-independent manner and that any potential de novo peripheral induction plays a minor role if any at least in the steady state in vivo.

gp130–STAT3-mediated Th17 development plays roles in autoimmune disease

It is important to evaluate the functional consequences of gp130–STAT3 pathway on Th17 development in vivo. CIA is reported to be a Th17-dependent autoimmune disease (13). We used F759 strain for evaluating the functional consequences of IL-6–gp130–STAT3-signaling pathway on Th17 development in vivo using this system. We established DBA background of F759 and control mice and then induced CIA. We showed that CIA was enhanced in F759 mice, whose gp130-mediated STAT3 signaling increased, compared with control animals (Fig. 3a). Moreover, serum IL-17 concentration increased in F759 mice compared with wild-type controls after induction of CIA (Fig. 3b). Thus, all the results presented here support the idea that Th17, which is easily induced in F759 mice, is at least partially involved in a Th17-mediated autoimmune disease, CIA-induced arthritis (13).

Fig. 3.

gp130–STAT3 pathway plays a role for development of CIA. (a) DBA background of F759 and control animals (males and females) were injected subcutaneously with CII with CFA (arrows: days 0 and 21). CIA development was enhanced in F759 mice compared with wild-type controls (males: P = 0.023, 0.017, 0.004, 0.002, 0.0006, 0.0005, 0.001 and 0.003 for days 28, 35, 39, 42, 46, 49, 53 and 56 after first immunization and females: P = 0.042, 0.024, 0.041, 0.017, 0.04 and 0.048 for days 28, 32, 35, 39, 53 and 56 after first immunization). (b) Serum IL-17 concentration was increased in F759 mice compared with controls at day 8 after second immunization (IL-17: P = 0.016).

In summary, the IL-6–gp130–STAT3-signaling pathway plays a role in autoimmune diseases at least in part through the induction of Th17 development with a minimum effect on Treg development at least in the steady state in vivo.

Supplementary data

Supplementary Figure 1 is available at International Immunology Online.

Abbreviations

Abbreviations

• APC
• CIA
  collagen-induced arthritis
• CII
• ERK
  extracellular signal-regulated kinase
• KO
• MAPK
  mitogen-activated protein kinase
• SHP2
  Src homology region 2 domain-containing phosphatase 2
• STAT
  signal transducers and activators of transcription
• TGFβ
  transforming growth factor β
• Treg

We thank S. Akira (Osaka University, Osaka, Japan) for the STAT3flox and lckCre mice. We are grateful to E. Iketani and T. Hayashi (Osaka University) for their excellent technical assistance and we thank R. Masuda (Osaka University) for her excellent secretarial assistance. This work was supported by a Grant-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology in Japan to K.I., M.M. and T.H. and from the Osaka Foundation for the Promotion of Clinical Immunology to M.M. and T.H.

References

1

TH1 and TH2 cells: different patterns of lymphokine secretion lead to different functional properties

,

Annu. Rev. Immunol.

,

1989

, vol.

7

pg.

145

2

The lineage decisions of helper T cells

,

Nat. Rev. Immunol.

,

2002

, vol.

2

pg.

933

3

Development of TH1 CD4+ T cells through IL-12 produced by _Listeria_-induced macrophages

,

Science

,

1993

, vol.

260

pg.

547

4

Natural killer cells are a source of interferon gamma that drives differentiation of CD4+ T cell subsets and induces early resistance to Leishmania major in mice

,

J. Exp. Med.

,

1993

, vol.

178

pg.

567

5

et al.

T-bet is a STAT1-induced regulator of IL-12R expression in naive CD4+ T cells

,

Nat. Immunol.

,

2002

, vol.

3

pg.

549

6

et al.

T-bet is rapidly induced by interferon-gamma in lymphoid and myeloid cells

,

Proc. Natl Acad. Sci. USA

,

2001

, vol.

98

pg.

15137

7

et al.

Basophils produce IL-4 and accumulate in tissues after infection with a Th2-inducing parasite

,

J. Exp. Med.

,

2004

, vol.

200

pg.

507

8

Helper T cells regulate type-2 innate immunity in vivo

,

Nature

,

2002

, vol.

420

pg.

825

9

Regulation of interleukin-12 signalling during T helper phenotype development

,

Adv. Exp. Med. Biol.

,

1996

, vol.

409

pg.

61

10

et al.

Interleukin 12 induces tyrosine phosphorylation and activation of STAT4 in human lymphocytes

,

Proc. Natl Acad. Sci. USA

,

1995

, vol.

92

pg.

7307

11

et al.

Lack of IL-4-induced Th2 response and IgE class switching in mice with disrupted Stat6 gene

,

Nature

,

1996

, vol.

380

pg.

630

12

et al.

Essential role of Stat6 in IL-4 signalling

,

Nature

,

1996

, vol.

380

pg.

627

13

Suppression of immune induction of collagen-induced arthritis in IL-17-deficient mice

,

J. Immunol.

,

2003

, vol.

171

pg.

6173

14

Interleukin-23 promotes a distinct CD4 T cell activation state characterized by the production of interleukin-17

,

J. Biol. Chem.

,

2002

, vol.

278

pg.

1910

15

et al.

Interleukin 17-producing CD4+ effector T cells develop via a lineage distinct from the T helper type 1 and 2 lineages

,

Nat. Immunol.

,

2005

, vol.

6

pg.

1123

16

et al.

IL-23 drives a pathogenic T cell population that induces autoimmune inflammation

,

J. Exp. Med.

,

2005

, vol.

201

pg.

233

17

et al.

A distinct lineage of CD4 T cells regulates tissue inflammation by producing interleukin 17

,

Nat. Immunol.

,

2005

, vol.

6

pg.

1133

18

et al.

Selective regulatory function of Socs3 in the formation of IL-17-secreting T cells

,

Proc. Natl Acad. Sci. USA

,

2006

, vol.

103

pg.

8137

19

et al.

Reciprocal developmental pathways for the generation of pathogenic effector TH17 and regulatory T cells

,

Nature

,

2006

, vol.

441

pg.

235

20

TGF in the context of an inflammatory cytokine milieu supports de novo differentiation of IL-17-producing T cells

,

Immunity

,

2006

, vol.

24

pg.

179

21

Signals mediated by transforming growth factor-beta initiate autoimmune encephalomyelitis, but chronic inflammation is needed to sustain disease

,

Nat. Immunol.

,

2006

, vol.

7

pg.

1151

22

et al.

The orphan nuclear receptor RORgammat directs the differentiation program of proinflammatory IL-17+ T helper cells

,

Cell

,

2006

, vol.

126

pg.

121

23

Tie2-Cre transgenic mice: a new model for endothelial cell-lineage analysis in vivo

,

Dev. Biol.

,

2001

, vol.

230

pg.

230

24

et al.

STAT3 regulates cytokine-mediated generation of inflammatory helper T cells

,

J. Biol. Chem.

,

2007

, vol.

282

pg.

9358

25

IL-6 signal transduction and its physiological roles: the signal orchestration model

,

Rev. Physiol. Biochem. Pharmacol.

,

2003

, vol.

149

pg.

1

26

Interleukin 6 and its receptor: ten years later

,

Int. Rev. Immunol.

,

1998

, vol.

16

pg.

249

27

et al.

Dissection of signaling cascades through gp130 in vivo: reciprocal roles for STAT3- and SHP2-mediated signals in immune responses

,

Immunity

,

2000

, vol.

12

pg.

95

28

et al.

A point mutation of Tyr-759 in interleukin 6 family cytokine receptor subunit gp130 causes autoimmune arthritis

,

J. Exp. Med.

,

2002

, vol.

196

pg.

979

29

et al.

Autoimmune arthritis associated with mutated interleukin (IL)-6 receptor gp130 is driven by STAT3/IL-7-dependent homeostatic proliferation of CD4+ T cells

,

J. Exp. Med.

,

2006

, vol.

203

pg.

1456

30

et al.

Postnatally induced inactivation of gp130 in mice results in neurological, cardiac, hematopoietic, immunological, hepatic, and pulmonary defects

,

J. Exp. Med.

,

1998

, vol.

188

pg.

1955

31

Stat3 activation is responsible for IL-6-dependent T cell proliferation through preventing apoptosis: generation and characterization of T cell-specific Stat3-deficient mice

,

J. Immunol.

,

1998

, vol.

161

pg.

4652

32

et al.

IL-6 regulates in vivo dendritic cell differentiation through STAT3 activation

,

J. Immunol.

,

2004

, vol.

173

pg.

3844

33

et al.

Pleiotropic defects of IL-6-deficient mice including early hematopoiesis, T and B cell function, and acute phase responses

,

Ann. N. Y. Acad. Sci.

,

1995

, vol.

762

pg.

308

34

et al.

The point mutation of tyrosine 759 of the IL-6 family cytokine receptor gp130 synergizes with HTLV-1 pX in promoting rheumatoid arthritis-like arthritis

,

Int. Immunol.

,

2004

, vol.

16

pg.

455

35

et al.

Toll-like receptor-mediated regulation of zinc homeostasis influences dendritic cell function

,

Nat. Immunol.

,

2006

, vol.

7

pg.

971

36

Immunization against heterologous type II collagen induces arthritis in mice

,

Nature

,

1980

, vol.

283

pg.

666

37

et al.

Thymus and autoimmunity: production of CD25+CD4+ naturally anergic and suppressive T cells as a key function of the thymus in maintaining immunologic self-tolerance

,

J. Immunol.

,

1999

, vol.

162

pg.

5317

38

et al.

Thymic selection of CD4+CD25+ regulatory T cells induced by an agonist self-peptide

,

Nat. Immunol.

,

2001

, vol.

2

pg.

301

Author notes

*

These authors contributed equally to this study.

© The Japanese Society for Immunology. 2007. All rights reserved. For permissions, please e-mail: journals.permissions@oxfordjournals.org

Topic:

• steady state
• autoimmune diseases
• mitogen-activated protein kinases
• t-lymphocytes
• interleukin-6
• mice
• tyrosine
• regulatory t-lymphocytes
• stat3 protein

Advertisement intended for healthcare professionals

Citations

Views

Altmetric

Metrics

Total Views 3,734

2,275 Pageviews

1,459 PDF Downloads

Since 1/1/2017

Citations

201 Web of Science

×

Email alerts

Citing articles via

• Latest

• Most Read

• Most Cited

More from Oxford Academic

Advertisement intended for healthcare professionals