| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
* Intractable Immune System Disease Research Center, Tokyo Medical University, Shinjuku, Tokyo, Japan;
Department of Patho-Physiology, Faculty of Pharmaceutical Sciences, Tokyo University of Science, Chiba, Japan;
Department of Immunology, Tokyo Medical University, Tokyo, Japan;
Department of Clinical Sciences, Faculty of Pharmaceutical Sciences, Josai International University, Chiba, Japan; and
¶ Department of Molecular Genetics, Medical Institute of Bioregulation, Kyushu University, Fukuoka, Japan
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionDisclosuresReferences |
|---|
production from naive CD4+ T cells stimulated with plate-coated anti-CD3 and anti-CD28 in the presence of IL-12, IL-27 simultaneously inhibited CD28-mediated IL-2 production. Correlated with the inhibition, IL-27 was shown to augment SOCS3 expression. Analyses using various mice lacking a signaling molecule revealed that the inhibition of IL-2 production was dependent on STAT1, but not on STAT3, STAT4, and T-bet, and was highly correlated with the induction of SOCS3 expression. Similar inhibition of CD28-mediated IL-2 production and augmentation of SOCS3 expression by IL-27 were observed in a T cell hybridoma cell line, 2B4. Forced expression of antisense SOCS3 or dominant negative SOCS3 in the T cell line blocked the IL-27-inudced inhibition of CD28-mediated IL-2 production. Furthermore, pretreatment with IL-27 inhibited IL-2-mediated cell proliferation and STAT5 activation, although IL-27 hardly affected the induction level of CD25 expression. These results suggest that IL-27 inhibits CD28-mediated IL-2 production and also IL-2 responses, and that SOCS3, whose expression is induced by IL-27, plays a critical role in the inhibitory effect in a negative feedback mechanism. |
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionDisclosuresReferences |
|---|
2 subunit, and gp130 constitute a functional signal-transducing receptor for IL-27 (1, 2). IL-27 activates JAK1, JAK2, TYK2, STAT1, STAT2, STAT3, STAT4, and STAT5 in naive CD4+ T cells (3, 4, 5, 6) and enhances proliferation in naive, but not memory, CD4+ T cells. IL-27 also induces the expression of T-bet, a master transcriptional regulator for Th1 differentiation (7), and subsequent IL-12R2 and synergizes with IL-12 in primary IFN- production (1, 3, 4, 6).
Previous studies using mice lacking one subunit of IL-27R, TCCR (8)/WSX-1 (9), revealed that IL-27 is required for the early initiation of Th1 responses, and that WSX-1/TCCR-deficient mice have enhanced susceptibility to infection with intracellular pathogens such as Leishmania major (9, 10) and Listeria monocytogenes (8). However, WSX-1 is not essential to develop the protective Th1 responses against Toxoplasma gondii parasites, but, rather, acts to attenuate the inflammatory responses induced by the protozoan infection, including cellular hyperactivation and overproduction of proinflammatory cytokines such as IFN-, IL-4, TNF-
, and IL-6 (5). In vitro analyses of the effect of IL-27 on Th1/Th2 differentiation demonstrated that IL-27 is not able to synergize with IL-12 to increase the production of IFN- by Th1 cells (11). Recent analyses of in vitro Th1/Th2 differentiation have revealed that the ability of IL-27 to induce Th1 differentiation is most prominent under Th1-polarizing conditions, but without IL-12, and is overruled by IL-12 (12). The IL-27-induced Th1 differentiation is mainly mediated by rapid and marked up-regulation of ICAM-1 expression on naive CD4+ T cells through ICAM-1/LFA-1 interaction in a STAT1-dependent, but T-bet-, IFN--, and STAT4-independent, mechanism. In contrast, it was recently demonstrated that IL-27 inhibits in vitro production of TNF and IL-12p40 in activated peritoneal macrophages from WSX-1+/+ mice, but not from WSX-1–/– mice. Taken together, these in vivo and in vitro results indicate that IL-27 not only plays a role in the early regulation of Th1 differentiation, but also exerts an inhibitory effect on immune responses, including the suppression of proinflammatory cytokine production. However, the molecular mechanism by which IL-27 exerts the inhibitory effect remains unclear.
In the present study, we have found that IL-27 inhibits CD28-mediated IL-2 production in CD4+ T cells and also IL-2 responses, and that suppressor of cytokine signaling (SOCS3), whose expression is induced by IL-27, mediates the inhibitory effect. Thus, IL-27, which is rapidly produced from APC by the interaction with T cells in the presence of Ag through CD40/CD40L interaction (1), plays important roles not only to augment T cell proliferation by itself and regulate early Th1 differentiation, but also to suppress excessive progression of CD28-mediated IL-2 production and IL-2 responses by inducing SOCS3 expression in a negative feedback mechanism.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionDisclosuresReferences |
|---|
BALB/c mice were purchased from Japan SLC. Mice transgenic (Tg) for TCR recognizing OVA323–339 (DO11.10; BALB/c background) (13) were provided by Dr. T. Yoshimoto (Hyogo College of Medicine, Hyogo, Japan). STAT1+/– and STAT1–/– mice (14) of a mixed background of 129/Sv and C57BL/6 were provided by Dr. R. D. Schreiber (Washington University, St. Louis, MO). STAT1-deficient mice (14) of 129/Sv background and wild-type 129/Sv mice were purchased from Taconic Farms. Mice lacking STAT3 specifically in T cells (Lck-Cre/STAT3flox/flox) were generated by mating STAT3flox/flox mice (15), in which the STAT3 gene is flanked by two loxP sites, and Lck-Cre Tg mice (16) (purchased from Center for Animal Resources and Development), in which the Cre recombinase transgene is regulated by T cell-specific Lck promoter. STAT3flox/flox, Lck-Cre/STAT3flox/+, or Lck-Cre/STAT3+/+ mice were used as control mice. STAT4-deficient mice (17) and T-bet-deficient mice (18) of BALB/c background were purchased from The Jackson Laboratory. All animal experiments were performed in accordance with our institutional guidelines.
Cells
Naive CD4+ T cells and a mouse T cell hybridoma cell line 2B4, provided by Dr. T. Saito (RIKEN Research Center for Allergy and Immunology, Kanagawa, Japan), were cultured in RPMI 1640 medium supplemented with 10% FBS and 50 µM 2-ME. PLAT-E, a packaging cell line provided by Dr. T. Kitamura (University of Tokyo, Tokyo, Japan) (19), was maintained in DMEM supplemented with 10% FBS.
Reagents
Anti-CD3 (145-2C11), anti-IL-2 (S4B6), anti-IL-4 (11B11), anti-IFN- (XMG1.2), and anti-Thy1.2 (30-H12) were purchased from American Type Culture Collection. Anti-CD28 (37.51) and mouse rIL-2 were obtained from BD Biosciences. Anti-CD25 (PC61.5) and FITC-anti-rat IgG were obtained from eBioscience. Anti-STAT1, anti-STAT3, anti-STAT5, and T-bet were purchased from Santa Cruz Biotechnology. Anti-phosphotyrosine (anti-pY)-STAT1, anti-pY-STAT3, and anti-pY-STAT5 were obtained from Cell Signaling Technology. Anti-SOCS3 and anti-actin were purchased from Medical Biological Laboratories and Sigma-Aldrich, respectively. Mouse rIL-12 was obtained from R&D Systems. Human rIL-2 and mouse rIFN- were provided by Shionogi.
Preparation of purified rIL-27 protein
Recombinant IL-27 was prepared as a soluble tagged fusion protein by flexibly linking EBV-induced gene 3 to p28 as described previously (20).
Preparation of naive CD4+ T cells
Primary T cells were purified by passing spleen cells depleted of erythrocytes through nylon wool. The flow-through fraction was incubated with biotin-conjugated anti-CD8, anti-B220, anti-Mac-1, anti-Ter-119, and anti-DX5, followed by incubation with anti-biotin magnetic beads (Miltenyi Biotec) and passed through a magnetic cell sorting column (Miltenyi Biotec); the negative fraction was collected (CD4+ T cells, >95%). These purified T cells were then incubated with anti-CD62L magnetic beads (Miltenyi Biotec), and the positive fraction was collected as purified naive CD4+ T cells (CD62L+ cells, >99%).
IL-2 and IFN- production assays
Naive CD4+ T cells (1 x 105 cells/ml) from DO11.10 Tg mice were stimulated with 1 µM OVA323–339 peptide and irradiated T/NK cell-depleted BALB/c spleen cells (1 x 106 cells/ml) in the presence or the absence of IL-27 (10 ng/ml) for various times. T/NK cell-depleted spleen cells were prepared as follows. Spleen cells depleted of erythrocytes were incubated with anti-Thy1.2, followed by incubation with anti-rat IgG magnetic beads (Miltenyi Biotec) together with anti-DX5 magnetic beads (Miltenyi Biotec) and passed through a magnetic cell-sorting column. The negative fraction was used as T/NK cell-depleted spleen cells. Naive CD4+ T cells (5 x 105 cells/ml) were stimulated with plate-coated anti-CD3 (2 µg/ml) and anti-CD28 (0.5 µg/ml) in the presence or the absence of IL-27 and/or IL-12 (10 ng/ml) for various times. 2B4 cells (2 x 105 cells/ml) were stimulated with plate-coated anti-CD3 (0.03 µg/ml) and anti-CD28 (0.5 µg/ml) in the presence or the absence of IL-27 for 16 h. Culture supernatant was collected and analyzed for IL-2 and/or IFN- production by ELISA (21).
RT-PCR analysis
Total RNA was extracted by using a guanidine thiocyanate procedure, cDNA was prepared using oligo(dT) primer and SuperScript reverse transcriptase (Invitrogen Life Technologies), and RT-PCR was performed using Taq DNA polymerase as described previously (22). Cycle conditions were 94°C for 40 s, 60°C for 20 s, and 72°C for 40 s. Primers used for hypoxanthine phosphoribosyltransferase (HPRT) were described previously (23). The following primers were also used; SOCS3 sense primer, 5'-TTGTCGGAAGACTGTCAACG-3'; SOCS3 antisense primer, 5'-GAGAGTCCGCTTGTCAAAGG-3'; WSX-1 sense primer, 5'-ACCCAAATGAAGCCAGACAC-3'; WSX-1 antisense primer, 5'-CACACAAGGTCTTGGGTCCT-3'; gp130 sense primer, 5'-AGTCTGGGTGGAAGCAGAGA-3'; and gp130 antisense primer, 5'-CTTGGTGGTCTGGATGGTCT-3'.
Quantitative RT-PCR analysis
cDNA synthesis was performed as described above. Real-time PCR was performed on an ABI 7500 (Applied Biosystems). PCR primers and probes for mouse SOCS3 and HPRT in the TaqMan Rodent Control Reagents and TaqMan Gene Expression Assays (Applied Biosystems), respectively, were used according to the manufacturer’s instructions. PCR parameters are as recommended for the TaqMan Universal PCR Master Mix kit (Applied Biosystems). Triplicate samples of 2-fold serial dilutions of cDNA were assayed and used to construct the standard curves.
Preparation of 2B4 transfectants
SOCS3 cDNA was isolated by RT-PCR using total RNA prepared from Con A-activated spleen cells and was confirmed by sequencing. Antisense SOCS3 cDNA (24) was generated using standard PCR methods and subcloned into p3xFLAG-CMV-10 vector (Sigma-Aldrich). 2B4 cells were then transfected with the antisense SOCS3 expression vector or the empty vector as a control by electroporation and selected with geneticin (G418).
Retroviral infection
Wild-type SOCS3, dominant-negative SOCS3(F25A) (25, 26), and antisense SOCS3 cDNAs were generated using standard PCR methods and subcloned into a bicistronic retroviral vector pMX-IRES/EGFP (27), provided by Dr. T. Kitamura. The PLAT-E cell line was transfected with the resultant vectors or the empty vector as control by using FuGene 6 (Roche) and cultured to generate the retroviral supernatant. 2B4 cells were then infected with the supernatant as described previously (28) and purified by sorting using a FACSVantage (BD Biosciences).
Proliferation assay
Naive CD4+ T cells (1 x 106 cells/ml) were stimulated with plate-coated anti-CD3 (2 µg/ml) and anti-CD28 (0.5 µg/ml) in the presence or the absence of IL-27. After 3 days, stimulated cells were recovered and washed. Resultant cells (1 x 105 cells/ml) were cultured in the presence of human IL-2 for 48 h and pulsed with [3H]thymidine for the last 24 h.
Western blotting
Cells were lysed in a lysis buffer containing protease inhibitors, and resultant cell lysates were separated by SDS-PAGE under reducing conditions and transferred to polyvinylidene difluoride membrane (Millipore) as described previously (28). The membrane was blocked, probed with primary Ab and then with the appropriate secondary Ab conjugated to HRP, and visualized with the ECL detection system (Amersham Biosciences) according to the manufacturer’s instructions.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionDisclosuresReferences |
|---|
We and other groups previously reported that IL-27 induces T-bet and subsequent IL-122 expression in naive CD4+ T cells and synergizes with IL-12 in IFN- production (1, 3, 4, 6). Although T-bet transcriptionally up-regulates IFN- production, it was originally demonstrated to down-regulate IL-2 production as well (7). In addition, WSX-1-deficient CD4+ T cells were reported to overproduce IL-2 (5). Therefore, we first examined the effect of IL-27 on primary IL-2 production. Naive CD4+ T cells from DO11.10 Tg mice were stimulated with OVA323–339 peptide and irradiated T/NK cell-depleted BALB/c spleen cells in the presence or the absence of IL-27 (10 ng/ml) for various times, and culture supernatant was collected and analyzed for IL-2 production by ELISA. The Ag-specific IL-2 production in naive CD4+ T cells gradually increased with time, and IL-27 greatly inhibited IL-2 production (Fig. 1A). Furthermore, naive CD4+ T cells from wild-type BALB/c mice were stimulated with plate-coated anti-CD3 and anti-CD28 in the presence of IL-27 and/or IL-12 for various times and analyzed for IL-2 and IFN- production (Fig. 1, B and C). As reported previously (1, 3, 4, 6), IL-27 induced T-bet expression (data not shown) and synergistic IFN- production with IL-12 (Fig. 1C). In marked contrast, IL-27 clearly inhibited IL-2 production in a dose-dependent manner, whereas IL-12 failed to affect IL-2 production, but appeared to enhance the inhibitory effect of IL-27 (Fig. 1, B and D). Without costimulation by anti-CD28, IL-2 production was not detected under the experimental conditions (Fig. 1E). In the presence of anti-CD28, greater production of IL-2 was observed with the higher dose of anti-CD3 used for plate coating. IL-27 inhibited IL-2 production independently of the dose of anti-CD3. These results suggest that IL-27 inhibits CD28-mediated IL-2 production in naive CD4+ T cells stimulated with Ag plus APC and also with plate-coated anti-CD3 plus anti-CD28.
|
Recently, it was reported that the expression of SOCS3 in early T cell activation influences the ability of IL-2 production mediated by CD28 costimulation (25). In addition, IFN- and IL-6 activate STAT1 and STAT3, respectively, both of which lead to SOCS3 induction. Therefore, we next examined the effect of IL-27 on SOCS3 expression. Naive CD4+ T cells were stimulated with plate-coated anti-CD3 and anti-CD28 in the presence or the absence of IL-27 for various times, total RNA was prepared and analyzed for SOCS3 and HPRT mRNA expression by RT-PCR and real-time PCR (Fig. 2). As reported previously (25), primary unstimulated CD4+ T cells expressed a significant level of SOCS3 mRNA, and the expression rapidly decreased after stimulation. However, in the presence of IL-27, SOCS3 expression was quickly recovered and increased; this pattern appeared to correlate with the inhibition of IL-2 production (Fig. 1B). These results suggest that IL-27 induces SOCS3 expression in naive CD4+ T cells stimulated with plate-coated anti-CD3 and anti-CD28, implying that SOCS3 may play a role in the exertion of inhibitory effects by IL-27.
|
To further explore the correlation between inhibition of IL-2 production and induction of SOCS3 expression by IL-27 and also which IL-27 downstream signaling molecule is required for the inhibition of CD28-mediated IL-2 production, we next used various mice lacking each of these signaling molecules, T-bet, STAT1, STAT3, and STAT4. Naive CD4+ T cells were prepared from these knockout mice and respective control mice and stimulated with plate-coated anti-CD3 and anti-CD28 in the presence or the absence of IL-27. After 48 h, culture supernatant was collected and analyzed for IL-2 production by ELISA. Total RNA was also prepared and analyzed for SOCS3 and HPRT mRNA expression by RT-PCR. Inhibition of IL-2 production by IL-27 was still observed in STAT3-, STAT4-, and T-bet-deficient naive CD4+ T (Fig. 3, B–D). However, in STAT1–/– naive CD4+ T cells, IL-2 production was hardly inhibited by IL-27 compared with that in STAT1+/– naive CD4+ T cells (Fig. 3A). Consistent with these results, the induction of SCOS3 mRNA expression was still observed in STAT3-, STAT4-, and T-bet-deficient naive CD4+ T cells, but not in STAT1-deficient naive CD4+ T cells (Fig. 3). These results suggest that the inhibition of IL-2 production is highly correlated with the induction of SOCS3 expression, and that STAT1 is required for inhibition of CD28-mediated IL-2 production and induction of SOCS3 expression by IL-27, although STAT3, STAT4, and T-bet are not essential to them.
|
To clarify a role for SOCS3 in the inhibition of CD28-mediated IL-2 production by IL-27, we next used a CD4+ T cell hybridoma cell line, 2B4, instead of primary naive CD4+ T cells. We first confirmed that 2B4 cells express both IL-27R subunits, gp130 and WSX-1, which were determined by RT-PCR (data not shown), and that 2B4 cells are responsive to IL-27, resulting in activation of STAT1 and STAT3, which was detected by Western blotting using anti-pY-STAT1 and anti-pY-STAT3 (data not shown). Then, 2B4 cells were stimulated with plate-coated anti-CD3 and anti-CD28 in the presence of IL-27. Culture supernatant was collected after 16 h and analyzed for IL-2 production by ELISA (Fig. 4A). Although stimulation with plate-coated anti-CD3 alone induced the production of significant amounts of IL-2, the addition of anti-CD28 further enhanced IL-2 production. Consistent with the results obtained using primary naive CD4+ T cells (Fig. 1D), IL-27 efficiently inhibited the CD28-mediated IL-2 production to the level obtained after stimulation with plate-coated anti-CD3 alone in a dose-dependent manner. Moreover, total RNA was prepared after the 3-h stimulation and analyzed for mRNA expression of SOCS3 by RT-PCR and real-time PCR (Fig. 4B). Correlated with the inhibition of IL-2 production, SOCS3 mRNA expression was induced in the presence of IL-27. Thus, the inhibition of CD28-mediated IL-2 production and the induction of SOCS3 expression by IL-27 were observed not only in primary naive CD4+ T cells, but also in a T cell hybridoma cell line, 2B4.
|
Because a T cell line is more suitable for gene transduction, we next generated 2B4 cells devoid of functional SOCS3 expression by transducing antisense SOCS3 or dominant negative SOCS3(F25A). We first prepared 2B4 cells expressing antisense SOCS3 and its empty vector as a control by transfecting and selection with geneticin (G418). Resultant stable transfectants (two clones each) were then stimulated with plate-coated anti-CD3 and anti-CD28 in the presence or the absence of IL-27. After stimulation for 3 h, total RNA was prepared and analyzed for mRNA expression of SOCS3 by RT-PCR. Augmentation of SOCS3 mRNA expression by IL-27 was barely observed in 2B4 transfectants expressing antisense SOCS3, although augmentation was clearly observed in 2B4 transfectants expressing control vector (Fig. 5A). Correlated with the inability to augment SOCS3 mRNA expression, inhibition of IL-2 production by IL-27 was not detected in 2B4 transfectants expressing antisense SOCS3, although the inhibition was clearly detected in 2B4 transfectants expressing control vector (Fig. 5A).
|
Taken together, these results suggest that IL-27 inhibits CD28-mediated IL-2 production, and that SOCS3, whose expression is induced by IL-27, plays a critical role in the inhibitory effect.
IL-27 inhibits IL-2-mediated cell proliferation and STAT5 activation without affecting CD25 expression
When naive CD4+ T cells were stimulated with plate-coated anti-CD3 and anti-CD28 in the presence or the absence of IL-27 for 3 days and then expanded in medium containing IL-2 for 3 more days, we initially noticed that recovery of the cell number in the presence of IL-27 appears to be less than that in the absence of IL-27 (data not shown). This implies that IL-27 may affect IL-2-mediated cell proliferation in addition to IL-2 production. Therefore, we finally investigated the effect of pretreatment with IL-27 on IL-2 responses. Naive CD4+ T cells were stimulated with plate-coated anti-CD3 and anti-CD28 in the presence or the absence of IL-27 for 3 days, and then these cells were washed and analyzed for the responsiveness to IL-2 by determining IL-2-mediated cell proliferation and tyrosine phosphorylation of STAT5, which is a critical signaling molecule activated by IL-2 (Fig. 6, A and B, respectively). Naive CD4+ T cells stimulated in the absence of IL-27 efficiently proliferated in response to increasing amounts of IL-2. In contrast, pretreatment with IL-27 reduced IL-2-mediated cell proliferation dose-dependently. The pretreatment also inhibited IL-2-induced STAT5 phosphorylation in STAT1+/+ naive CD4+ T cells, but not in STAT1–/– naive CD4+ T cells. This is consistent with the finding that STAT1 is required for induction of SOCS3 expression by IL-27 (Fig. 3). We then examined the effect of IL-27 on the induction of CD25 (IL-2R) expression by FACS analysis. Stimulation with plate-coated anti-CD3 and anti-CD28 greatly enhanced CD25 expression on naive CD4+ T cells, whereas comparable induction of CD25 expression was observed in the presence and the absence of IL-27 (Fig. 6C). These results suggest that IL-27 inhibits IL-2-mediated cell proliferation and STAT5 activation without affecting CD25 expression as well as IL-2 production.
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionDisclosuresReferences |
|---|
gene and a master transcriptional regulator for Th1 differentiation, whereas it simultaneously represses IL-2 gene transcription (7). IL-27 can augment T-bet and subsequent IL-12R2 expression in naive CD4+ T cells, resulting in synergistic IFN- production with IL-12 (1, 3, 4, 6). Therefore, we initially expected that T-bet might be required for the inhibition of IL-2 production by IL-27. However, it turned out that SOCS3, but not T-bet, is required for the inhibition of IL-2 production (Fig. 3). Moreover, it was recently demonstrated that SOCS3 expression induced by IFN- is achieved via activation of STAT1, but not STAT3 (29). Similarly, IL-27 was revealed to induce SOCS3 expression via activation of STAT1, but not STAT3 (Fig. 3), although IL-27 can activate both STAT1 and STAT3 efficiently (3, 4, 5, 6). Previously, it was demonstrated that SOCS3 is rapidly induced by IL-2 in T cells and inhibits IL-2 responses, including STAT5 phosphorylation and proliferation, in a negative feedback loop (30). It was also demonstrated that Ag stimulation of naive CD4+ T cells down-regulates SOCS3 expression, which is subsequently followed by a gradual increase in the SOCS3 level and corresponding decline in IL-2 secretion (24). Moreover, forced overexpression of SOCS3 inhibits proliferation of CD4+ T cells, whereas depletion of endogenous SOCS3 by antisense SOCS3 cDNA enhances TCR- and cytokine-induced proliferation. Thus, SOCS3 has antagonistic effects on IL-2 production and IL-2 responses, resulting in a feedback regulation of T cell activation. Recently, SOCS3 has been also shown to play a role in prohibiting excessive progression of CD28-mediated IL-2 production in a negative feedback mechanism (25). SOCS3 is expressed in the primary IL-2 production process and then interacts with phosphorylated CD28 through its SH2 domain by competing with PI3K, whose binding to the phosphorylated CD28 is considered to be crucial for CD28-mediated IL-2 production. Taken together with the present finding that IL-27 suppresses CD28-mediated IL-2 production through SOCS3 induction, it is highly conceivable that IL-27, which is rapidly produced from APC by the interaction with T cells in the presence of Ag through CD40/CD40L interaction (1), has important roles not only to augment T cell proliferation by itself and regulate early Th1 differentiation, but also to suppress excessive progression of CD28-mediated IL-2 production and IL-2-mediated cell proliferation by inducing SOCS3 expression in a negative feedback mechanism.
IFN- is a pluripotent cytokine that has a crucial role in several processes, including host defense against viruses and microorganisms, antiproliferative effect, phagocyte activation, control of apoptosis, promotion of Ag processing and presentation, and Th1 differentiation (31). STAT1 plays a major role in mediating these actions by IFN-, although STAT3 and STAT5 can also be activated by IFN- in certain cell types. IL-6 is also a multifunctional cytokine that regulates inflammatory responses, hemopoiesis, and the acute phase response (32). Greater production of IL-6 is associated with immune-mediated diseases, such as rheumatoid arthritis. IL-6 activates STAT1 and STAT3, whereas gene deletion studies have indicated that STAT3 has a primary role in determining the cellular responses. Although STAT1 and STAT3 are very similar proteins often activated by the same stimuli, they have very different effects on cell growth and survival. It is considered that STAT1 is a tumor suppressor (33, 34), whereas STAT3 is an oncogene (35). The reciprocal activation of these two transcriptional factors in response to IFN- or IL-6 suggests that their relative abundance, which may vary substantially in different cell types, under different conditions is likely to have a major impact on how cells behave in response to these two cytokines. Because IL-27 efficiently activates both STAT1 and STAT3 presumably through different IL-27R subunits, WSX-1 and gp130, respectively (3, 4, 5, 6), physiological consequences in response to IL-27 could be more complicated and determined in the balance between STAT1 and STAT3 activation and also their negative feedback inhibitors, SOCS3 as shown in the present study and presumably SOCS1, whose role in IL-27-mediated immune responses remains to be elucidated.
|
Acknowledgments |
|---|
|
Disclosures |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionDisclosuresReferences |
|---|
|
Footnotes |
|---|
1 This study was supported by Grant-in-Aid for Scientific Research, High-Tech Research Center Project, and University-Industry Joint Research Project from the Ministry of Education, Culture, Sports, Science, and Technology of Japan and a grant from Novartis Foundation (Japan) for the Promotion of Science. 
2 Address correspondence and reprint requests to Dr. Takayuki Yoshimoto, Intractable Immune System Disease Research Center, Tokyo Medical University, 6-1-1 Shinjuku, Shinjuku-ku, Tokyo 160-8402, Japan. E-mail address: yoshimot{at}tokyo-med.ac.jp
3 Abbreviations used in this paper: TCCR, T cell cytokine receptor; HPRT, hypoxanthine phosphoribosyltransferase; pY, phosphotyrosine; SOCS, suppressor of cytokine signaling; Tg, transgenic.
Received for publication July 13, 2005. Accepted for publication December 29, 2005.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionDisclosuresReferences |
|---|
signal via Stat1 and Stat3 and induce T-Bet and IL-12R2 in naive T cells. J. Interferon Cytokine Res. 23: 513-522. [Medline] production in CD4 and CD8 T cells. Science 295: 338-342. 1 cytoplasmic region. J. Immunol. 171: 1352-1359. . J. Biol. Chem. 279: 13746-13754. receptor: a paradigm for cytokine receptor signaling. Annu. Rev. Immunol. 15: 563-591. [Medline] and interferon . Proc. Natl. Acad. Sci. USA 93: 7673-7678. This article has been cited by other articles:
|
|
 |
|
  T. Yoshimoto, N. Morishima, I. Mizoguchi, M. Shimizu, H. Nagai, S. Oniki, M. Oka, C. Nishigori, and J. Mizuguchi Antiproliferative Activity of IL-27 on Melanoma J. Immunol., May 15, 2008; 180(10): 6527 - 6535. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Owaki, M. Asakawa, N. Morishima, I. Mizoguchi, F. Fukai, K. Takeda, J. Mizuguchi, and T. Yoshimoto STAT3 Is Indispensable to IL-27-Mediated Cell Proliferation but Not to IL-27-Induced Th1 Differentiation and Suppression of Proinflammatory Cytokine Production J. Immunol., March 1, 2008; 180(5): 2903 - 2911. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Seita, M. Asakawa, J. Ooehara, S.-i. Takayanagi, Y. Morita, N. Watanabe, K. Fujita, M. Kudo, J. Mizuguchi, H. Ema, et al. Interleukin-27 directly induces differentiation in hematopoietic stem cells Blood, February 15, 2008; 111(4): 1903 - 1912. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Huber, V. Steinwald, A. Guralnik, A. Brustle, P. Kleemann, C. Rosenplanter, T. Decker, and M. Lohoff IL-27 inhibits the development of regulatory T cells via STAT3 Int. Immunol., February 1, 2008; 20(2): 223 - 234. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Cao, P. D. Doodes, T. T. Glant, and A. Finnegan IL-27 Induces a Th1 Immune Response and Susceptibility to Experimental Arthritis J. Immunol., January 15, 2008; 180(2): 922 - 930. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Wang, Y. Miyazaki, Y. Shinozaki, and H. Yoshida Augmentation of Antigen-Presenting and Th1-Promoting Functions of Dendritic Cells by WSX-1(IL-27R) Deficiency J. Immunol., November 15, 2007; 179(10): 6421 - 6428. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Pirhonen, J. Siren, I. Julkunen, and S. Matikainen IFN-{alpha} regulates Toll-like receptor-mediated IL-27 gene expression in human macrophages J. Leukoc. Biol., November 1, 2007; 82(5): 1185 - 1192. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. Qin, K. L. Roberts, S. A. Niyongere, Y. Cong, C. O. Elson, and E. N. Benveniste Molecular Mechanism of Lipopolysaccharide-Induced SOCS-3 Gene Expression in Macrophages and Microglia J. Immunol., November 1, 2007; 179(9): 5966 - 5976. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Brender, G. M. Tannahill, B. J. Jenkins, J. Fletcher, R. Columbus, C. J. M. Saris, M. Ernst, N. A. Nicola, D. J. Hilton, W. S. Alexander, et al. Suppressor of cytokine signaling 3 regulates CD8 T-cell proliferation by inhibition of interleukins 6 and 27 Blood, October 1, 2007; 110(7): 2528 - 2536. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. C. Fitzgerald, B. Ciric, T. Touil, H. Harle, J. Grammatikopolou, J. D. Sarma, B. Gran, G.-X. Zhang, and A. Rostami Suppressive Effect of IL-27 on Encephalitogenic Th17 Cells and the Effector Phase of Experimental Autoimmune Encephalomyelitis J. Immunol., September 1, 2007; 179(5): 3268 - 3275. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Liu, X. Guan, and X. Ma Regulation of IL-27 p28 gene expression in macrophages through MyD88- and interferon-{gamma}-mediated pathways J. Exp. Med., January 22, 2007; 204(1): 141 - 152. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Owaki, M. Asakawa, F. Fukai, J. Mizuguchi, and T. Yoshimoto IL-27 Induces Th1 Differentiation via p38 MAPK/T-bet- and Intercellular Adhesion Molecule-1/LFA-1/ERK1/2-Dependent Pathways J. Immunol., December 1, 2006; 177(11): 7579 - 7587. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Yoshimura, A. Takeda, S. Hamano, Y. Miyazaki, I. Kinjyo, T. Ishibashi, A. Yoshimura, and H. Yoshida Two-Sided Roles of IL-27: Induction of Th1 Differentiation on Naive CD4+ T Cells versus Suppression of Proinflammatory Cytokine Production Including IL-23-Induced IL-17 on Activated CD4+ T Cells Partially Through STAT3-Dependent Mechanism J. Immunol., October 15, 2006; 177(8): 5377 - 5385. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||
