The JI
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     
 

The Journal of Immunology, 2007, 178: 8138-8147.
Copyright © 2007 by The American Association of Immunologists, Inc.
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Sheibanie, A. F.
Right arrow Articles by Ganea, D.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Sheibanie, A. F.
Right arrow Articles by Ganea, D.

The Proinflammatory Effect of Prostaglandin E2 in Experimental Inflammatory Bowel Disease Is Mediated through the IL-23->IL-17 Axis1

Amir F. Sheibanie*, Jui-Hung Yen*,{dagger}, Tanzilya Khayrullina*,{dagger}, Frances Emig*, Ming Zhang*, Ronald Tuma* and Doina Ganea2,*

* Department of Physiology, Temple University School of Medicine, Philadelphia, PA 19140; and {dagger} Department of Biological Sciences, Rutgers University, Newark, NJ 07102


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
Although Crohn’s disease has been traditionally considered to be Th1-mediated, the newly identified Th17 cells emerged recently as crucial participants. Th1/Th17 differentiation is controlled primarily by the IL-12 family of cytokines secreted by activated dendritic cells (DCs) and macrophages. IL-23 and IL-12/IL-27 have opposite effects, supporting the Th17 and Th1 phenotypes, respectively. We found that PGE2, a major lipid mediator released in inflammatory conditions, shifts the IL-12/IL-23 balance in DCs in favor of IL-23, and propose that high levels of PGE2 exacerbate the inflammatory process in inflammatory bowel disease through the IL-23->IL-17 axis. We assessed the effects of PGE2 on IL-12, IL-27, and IL-23 and found that PGE2 promotes IL-23, inhibits IL-12 and IL-27 expression and release from stimulated DCs, and subsequently induces IL-17 production in activated T cells. The effects of PGE2 are mediated through the EP2/EP4 receptors on DCs. In vivo, we assessed the effects of PGE analogs in an experimental model for inflammatory bowel disease and found that the exacerbation of clinical symptoms and histopathology correlated with an increase in IL-23 and IL-17, a decrease in IL-12p35 expression in colon and mesenteric lymph nodes, and a substantial increase in the number of infiltrating neutrophils and of CD4+IL-17+ T cells in the colonic tissue. These studies suggest that high levels of PGE2 exacerbate the inflammatory process through the preferential expression and release of DC-derived IL-23 and the subsequent support of the autoreactive/inflammatory Th17 phenotype.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
Inflammatory bowel diseases (IBD),3 which include Crohn’s disease (CrD) and ulcerative colitis (UC), are chronic, relapsing disorders, characterized by inflammatory reactions to microbial Ags of the commensal flora (1, 2). Recent studies highlighted the importance of both adaptive and innate immunity in IBD pathogenesis (3, 4, 5).

CrD was traditionally considered a Th1-type disease (6, 7, 8). Recently however, the Th17 cells emerged as crucial mediators in autoimmune disorders, including IBD (9, 10, 11, 12, 13, 14, 15). CrD and UC patients, but not healthy individuals or patients with infectious or ischemic colitis, have increased IL-17 levels in serum and in the colonic mucosa (16, 17, 18). In animal models of colitis, the administration of anti-IL-17 Abs ameliorates intestinal inflammation (19). In addition, IL-17R-deficient mice are less susceptible to 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis, although they express higher levels of IFN-{gamma}, a hallmark of the Th1 response (20).

Members of the IL-12 cytokine family, i.e., IL-12, IL-23, and IL-27, are important in Th1/Th17 differentiation (21, 22, 23). In contrast to IL-23, which stabilizes the Th17 phenotype (14, 19, 24, 25, 26, 27), IL-12 and IL-27 promote Th1 responses (28, 29, 30, 31, 32) and suppress Th17 development and function (33, 34, 35). Therefore, the IL-12/IL-27 vs IL-23 balance has a direct effect on the ensuing numbers of Th1/Th17 effectors.

We reported previously that PGE2 induces IL-23 and inhibits IL-12p70 release from dendritic cells (DC) (36). In this study, we propose that high levels of PGE2 released locally in inflammatory/autoimmune conditions sustain inflammation by activating resident DC to express IL-23 and shifting the T cell response toward Th17. We addressed this hypothesis in vivo in the TNBS-induced colitis model which shares many characteristics with human CrD in terms of ulceration, inflammation, leukocyte infiltration, and transmural lesions (37).


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 Disclosures
 References
 
Mice

B10.A and BALB/c mice (males, 6–8 wk old; 20–22 g; The Jackson Laboratory) were maintained in the Temple University Animal Facility under pathogen-free conditions. The animal protocols were approved by the Committee on Use and Care of Laboratory Animals at Temple University (249-A3).

Reagents

TNBS, LPS (Escherichia coli 055:B5), and indomethacin were obtained from Sigma-Aldrich. The myeloperoxidase assay kit was obtained from CytoStore. Recombinant murine (rm) GM-CSF was purchased from PeproTech. Capture and biotinylated anti-mouse IL-17, rmIL-17, and the ELISA kit for murine IL-27p28 were obtained from R&D Systems. Capture and biotinylated anti-mouse IL-6, IL-1beta, IFN-{gamma}, TNF, IL-10, IL-12p40, IL-12p70, FITC-conjugated anti-CD4, PE-conjugated anti-IL-17 Abs, anti-CD3, and anti-CD28 mAbs were purchased from BD Pharmingen. The ELISA kit for murine IL-23 and rmIL-23 standards were obtained from eBioscience. PGE2, misoprostol, and PGE1-OH were purchased from Cayman Chemical. The FD Rapid Multistain kit used for H&E staining was purchased from FD Neurotechnologies.

Generation of bone marrow-derived DCs

DC were generated in vitro from B10.A bone marrow as previously described (38). Purified CD11c+ DC were cultured with LPS (1 µg/ml), PGE2 (10–5 or 10–6 M), or both, for 3 h for RNA preparation and 12 h for ELISA.

IL-17 production by activated T cells

CD4+ T cells were purified from the spleen of B10A mice using anti-CD4-coupled magnetic beads. The CD4+ T cells (>97% by FACS) were cultured at a concentration of 2 x 106 cells/ml in 24-well plates (250 µl/well) precoated with anti-CD3 mAbs (2.5 µg/well), and 4 days later the amount of secreted IL-17 was determined by ELISA. The detection limit for the IL-17 ELISA is 15 pg/ml.

The TNBS colitis model

Colitis was induced in BALB/c males (20–22 g) anesthetized with isoflurane, by intracolonic administration of TNBS (3 mg in 50% ethanol). Groups receiving misoprostol were injected i.p. with misoprostol (2, 20, or 60 µg/animal) or PGE1-OH (5 or 30 µg/animal) 1 h after TNBS, followed by another injection 12 h later. The mice were weighed every 12 h. Control groups received 50% ethanol or 50% ethanol plus the highest dose of misoprostol or PGE1-OH. Groups receiving indomethacin were injected i.p. with indomethacin (1.5 mg/kg) 30 min before TNBS administration. The mice received indomethacin (same dose, i.p.) every 12 h on days 1 and 2 and were weighed daily.

Assessment of inflammation

Each colon received a macroscopic score (from 0 to 10) by two independent blinded observers as described by Chin et al. (39). For histologic score, colon sections were stained with H&E and scored as follows: 0, no inflammation; 1, low leukocyte infiltration; 2, moderate leukocyte infiltration; 3, high infiltration, moderate fibrosis and goblet loss; and 4, massive loss of goblet cells, extensive fibrosis, and thickening of the colon wall.

Real-time RT-PCR

The SYBR green-based real-time PCR technique was used to detect the expression of p19, p35, p40, IL-17, and IFN-{gamma} as previously described (38) The specific primers were as follows: p19 forward (F) 5'-TGCTGGATTGCAGAGCAGTAA-3' and reverse (R) 5'-GCATGCAGAATTCCGAAGA-3'; p35 F 5'-GAGGACTTGAAGATGTACAG-3' and R 5'-TTCTACTTGGAGGAGGGC-3'; p40 F 5'-GACCCTGCCGATTGAACTGGC-3' and R 5'-CAACGTTGCATCCTAGGATCG-3'; GAPDH F 5'-GTTCAGCTCTGGATGACCTTGCC-3' and R 5'-TCCTGCACCACCAACTGCTTAGCC-3'; beta-actin F 5'-TCCACCACCACAGCTGAGAGG-3' and R 5'-CAGCTTCTCTTTGATGTCACG-3'; IL-17 F 5'-CCACACCCACAGCATCTC-3'; and R 5'-CTGAGAGCTGCCCCTTCAC-3'; IFN-{gamma} F 5'-GCTTCTGAGCTGATTCC-3' and R 5'-AGCTGATCCGAGTGGTCCAC-3'; p28 forward 5'-TCTGGTACAAGCTGGTTCCTGG-3' and R 5'-TAGCCCTGAACCCAGAAGA-3'; EBI3 F 5'-CACGGTGCCCTACATGCTAA-3' and R 5'-GAGGTCGGTTGATGATT-3'.

Measurement of myeloperoxidase (MPO) activity

A total of 50 mg/ml colon tissue was homogenized and the MPO activity was measured as described previously (40). MPO activity was expressed as units per gram of tissue.

Cytokine determination by ELISA

Supernatants collected from DC cultures and from mesenteric lymph nodes (MLN) harvested from mice with colitis and restimulated ex vivo (see below) were assayed for cytokines by ELISA. The detection limits are 15 pg/ml for IL-17, IL-6, IL-12p40, IL-4, IL-10, and IL-23, 30 pg/ml for IFN-{gamma} and IL-12p70, 7.8 pg/ml for IL-27p28, and 5 pg/ml for TNF. For measurement of tissue cytokines, colonic tissue was processed as previously described (40) and cytokine content was determined by ELISA.

Isolation and culture of MLN

MLN were collected 36 h after TNBS administration. Single-cell suspensions (1 x 106 cells/ml) were restimulated with immobilized anti-CD3 mAb (2.5 µg/ml) and soluble anti-CD28 mAb (2 µg/ml). Supernatants collected 72 h later were assayed for cytokines by ELISA.

Histology

Tissue samples were fixed and stained with H&E. For immunohistochemistry, the slides were incubated with 3% BSA for 30 min, washed, and incubated with PE-labeled anti-IL-17 (5 µg/ml) and FITC-labeled anti-CD4 (5 µg/ml) Abs. After incubation, the sections were covered with prolong anti-fade (Molecular Probes) and analyzed using a Nikon TE200 epifluorescence microscope, equipped with a Hamamatsu camera. Red filters (range 575–615 nm) and green filters (range 500–550 nm) were used for PE and FITC staining, respectively.

Statistical analysis

All values are expressed as mean ± SD. Survival curves were analyzed by Kaplan-Meier log-rank test. Changes in body weight were compared by Kruskal-Wallis ANOVA. The in vitro data were analyzed by two-tailed Student’s t test for unpaired samples and ANOVA for comparison of multiple groups. Macroscopic and histologic scores between TNBS and TNBS plus misoprostol groups were analyzed with a two-sided Wilcoxon rank-sum test.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 Disclosures
 References
 
PGE2 inhibits IL-12 and IL-27 and stimulates IL-23 expression in LPS-treated DC; role of EP receptors

We have shown previously that PGE2 induces IL-23 but not IL-12 release from immature DC and inhibits IL-12p70 production in LPS-stimulated DC (36). Because IL-12 (p40/p35) and IL-27 (p28/EBI3) induce Th1 responses, whereas IL-23 (p40/p19) is essential for the Th17 response, we determined the effects of PGE2 on p40, p35, p19, p28, and EBI3 expression in LPS-stimulated DC. In time-course experiments, we established that all subunits are expressed early, with maximum induction at 3 h (results not shown). Purified CD11c+ DC were treated with LPS with or without PGE2 and p40, p35, p19, p28, and EBI3 expression levels were determined 3 h later by RT-PCR. PGE2 inhibited p35, reduced p40, and increased p19 expression (Fig. 1A), shifting the IL-12/IL-23 balance in favor of IL-23. As for IL-27, the EBI3 subunit is constitutively expressed in DC and up-regulated by LPS, whereas the p28 subunit is induced following LPS stimulation (Fig. 1B). PGE2 does not affect EBI3 levels, but reduces p28 expression (Fig. 1B).


Figure 1
View larger version (30K):
[in this window]
[in a new window]

  		FIGURE 1. PGE2 promotes IL-23 and inhibits IL-12 and IL-27 production in LPS-stimulated DC. A–C, CD11c+ DC (2 x 106 cells/ml) were stimulated with LPS (1 µg/ml) with or without PGE2 (10–6 M). RNA was prepared 3 h later and subjected to RT-PCR with primers for p19, p40, and p35 (A), and for p28 and EBI3 (B). Supernatants were collected at 12 h and assayed by ELISA for IL-12p40, IL-12p70, IL-27p28, and IL-23 (C). The data are expressed as mean ± SD from four separate experiments. D, Upper panel, CD11c+ DC (2 x 106 cells/ml) were stimulated with LPS (1 µg/ml) and treated with PGE2 (10–6 M), sulprostone, butaprost, or misoprostol (10–5 M). RNA was prepared 3 h later and subjected to RT-PCR with primers for p19. Lower panel, CD11c+ DC (1 x 106 cells/ml) were cultured in the presence of PGE2 (10–6 M), sulprostone, butaprost, or misoprostol (10–5 M) for 24 h. Supernatants were collected and added to CD4+ T cells cultured in the presence of immobilized anti-CD3 mAbs for 72 h. IL-17 release in the supernatants was measured by ELISA. Controls consisted of PGE2, sulprostone, butaprost, or misoprostol added directly to activated T cells, and T cells cultured in the absence of anti-CD3 Abs. The data are expressed as mean ± SD from three separate experiments.

 
The effect of PGE2 was confirmed at the protein level. Supernatants collected 12 h after treatment with LPS and PGE2 were assayed for IL-23, IL-12p40, IL-12p70, and IL-27p28 by ELISA. LPS induced all four cytokines. PGE2 more than doubled IL-23 levels, while suppressing IL-12p70 and IL-27p28 production (Fig. 1C). These results confirm that PGE2 shifts the balance in favor of IL-23 in LPS-stimulated DC.

PGE2 acts through four receptor subtypes, i.e., EP1–4. Previously, we have shown that DC express high levels of EP2 and EP4, and that the PGE2 effects on p19 expression in immature DC and on TNF and CCL3/4 production in TLR-stimulated DC are mediated through the EP2/EP4 receptors (36, 38, 41). To evaluate the role of EP receptors in the up-regulation of p19 in LPS-stimulated DC, we compared the effects of PGE2 and of various receptor agonists. CD11c+ DC were stimulated with LPS in the presence of PGE2, butaprost (an EP2 agonist), misoprostol (EP3>EP4>EP1>EP2), or sulprostone (EP3, EP1) for 3 h and the expression of p19 was determined by RT-PCR. Similar to PGE2, butaprost and misoprostol, but not sulprostone, induced p19 expression (Fig. 1D, upper panel), indicating the involvement of the EP2/EP4 receptors.

Previously, we have shown that supernatants collected from DC treated with PGE2 induced IL-17 production in activated T cells and that this effect was mediated, at least partially, through IL-23 (36). In this study, we tested the effect of supernatants collected from DC treated with PGE2 or various EP receptor agonists on IL-17 production. Direct addition of EP agonists to activated T cells did not result in IL-17 release. In contrast, supernatants harvested from DC treated with PGE2, misoprostol or butaprost, but not sulprostone, induced IL-17 release (Fig. 1D, lower panel). This indicates that EP2/EP4 receptors mediate the effect of PGE2 in inducing DC that promote IL-17 production in T cells.

PGE2 agonists exacerbate TNBS colitis

We hypothesized that PGE2 shifts locally the IL-12/IL27 vs IL-23 balance in favor of IL-23, leading to the preferential accumulation of Th17 cells. Based on the in vitro data, we chose two stable PGE analogs, misoprostol, a potent, nonselective agonist (EP4, EP3>EP1>EP2), and the EP4 agonist PGE1-OH (42) for the in vivo studies. Colitis was induced in BALB/c mice by a single intracolonic administration of TNBS in 50% ethanol. Misoprostol (60 µg/animal) was injected i.p. 1 and 12 h after TNBS administration. Control groups consisted of ethanol and ethanol plus misoprostol (no TNBS). One hundred percent of the animals survived in the control groups with no significant weight loss over the observation period (10 days) (Fig. 2, A and B). In contrast, 40% of the mice in the TNBS group and 90% in the TNBS plus miso group succumbed by day 3 (Fig. 2A). There was significant weight loss in the TNBS plus miso-treated group as compared with the TNBS control (Fig. 2B). Misoprostol by itself does not affect survival or induce wasting (Fig. 2A). Similar to misoprostol, administration of PGE1-OH (30 µg/mouse) to mice treated with TNBS resulted in 80% mortality (Fig. 2C). Lower doses of misoprostol (6 and 20 µg/mouse) and of PGE1-OH (5 µg/mouse) had marginal effects.


Figure 2
View larger version (30K):
[in this window]
[in a new window]

  		FIGURE 2. Misoprostol exacerbates colitis and induces colonic expression of IL-23 and IL-17. A–C, BALB/c mice (n = 10) were treated with 50% EtOH, EtOH and misoprostol (60 µg/mouse), TNBS (3 mg/mouse), and TNBS and misoprostol as described in Materials and Methods. Survival (A) and body weight (B) were monitored in a blinded fashion. Similar treatment except for PGE1-OH (5 or 30 µg/mouse) instead of misoprostol (C). The Kaplan-Meier log-rank test was used for the survival curves and the Kruskal-Wallis test was used for changes in body weight. One experiment of three is shown. *, p < 0.05 (comparison between misoprostol plus TNBS and TNBS treatment). D, Macroscopic and histology scores were determined using colons harvested 36 h after TNBS administration. E, Expression of p19, p35, and p40 in colonic tissue (36 h) was analyzed individually by real-time RT-PCR. One representative experiment of four is shown. F, Expression of IFN-{gamma} and IL-17 in colonic tissue (36 h) using real-time RT-PCR. One representative experiment of three is shown.

 
Macroscopic analysis of colons obtained 36 h after TNBS administration indicated striking hyperemia, ulceration, and necrosis in mice treated with misoprostol (Fig. 2D, upper panel). The histology score was also higher in mice treated with misoprostol (Fig. 2D, lower panel). The analyses were performed at 36 h because most of the mice treated with misoprostol died 60 h after TNBS administration. In mice treated only with TNBS, 36 h represent an early time point (maximum effects are observed at 72 h), and therefore our macroscopic and histology scores for TNBS-treated mice are lower than those reported in the literature.

Misoprostol affects colonic p19, p35, p40, and IL-17 expression

Colons from mice sacrificed 36 h after TNBS administration were processed for RT-PCR. TNBS induces increases in both IL-23 and IL-12 expression, i.e., up-regulation of p35, p40, and p19 (Fig. 2E). As expected, there is increased IFN-{gamma} and IL-17 expression. In contrast, animals treated with TNBS plus misoprostol show impressive increases in p40 and p19, and a significant decrease in p35 expression (Fig. 2E). This confirms the shift in the IL-12/IL-23 balance observed in vitro in DC. As expected, the levels of colonic IL-17 expression in TNBS plus miso-treated mice were significantly higher compared with TNBS (Fig. 2F, lower panel). These results suggest that the PGE2 agonist induces the IL-23->IL-17 axis in colon, presumably through Th17 cells, in addition to the existing Th1-mediated IFN-{gamma} response.

Effect of misoprostol on cytokine production in colon

Colons were processed for cytokine determination. We detected proinflammatory cytokines, i.e., IL-12p40, IFN-{gamma}, IL-17, IL-6, TNF, and IL-1beta, in the colons of TNBS-treated mice. Animals inoculated with TNBS and misoprostol showed increased levels of IL-17, IL-12 p40, IL-1beta, IL-6, and TNF, but not IFN-{gamma} (Fig. 3A).


Figure 3
View larger version (24K):
[in this window]
[in a new window]

  		FIGURE 3. Misoprostol induces colonic expression of proinflammatory cytokines and neutrophil infiltration. A, BALB/c mice (n = 7) were treated as described in Fig. 2. Colonic protein extracts (36 h) were subjected to ELISA for various cytokines. Data are represented as the mean ± SD with individual samples tested in triplicates. One experiment of four is shown. B, Cell suspensions from MLN harvested at 36 h (1 x 106 cells/ml) were restimulated ex vivo with immobilized anti-CD3 Abs (2.5 µg/ml) and soluble anti-CD28 Abs (2 µg/ml) for 72 h. Supernatants were assayed for IL-17, IFN-{gamma}, and IL-4 by ELISA. Data are represented as the mean ± SD. One experiment of three is shown. C, Intestinal protein extracts (36 h) were subjected to MPO determination. Data are presented as mean ± SD. One experiment of three is shown.

 
Effect of misoprostol on IL-17, IFN-{gamma}, and IL-4 production in MLN

MLN cells were restimulated ex vivo with anti-CD3 and anti-CD28 mAb. Supernatants collected 72 h later were subjected to ELISAs for IL-17, IFN-{gamma}, and IL-4. Cells from mice inoculated with TNBS plus miso released more IL-17 and less IFN-{gamma} than cells from mice inoculated with TNBS alone (Fig. 3B). There was no significant IL-4 production in any of the four groups. Taken together with the p19/p40/p35 expression data, these results suggest that misoprostol promotes Th17, but not Th1 and Th2 function, presumably through preferential induction of IL-23.

Misoprostol increases neutrophil infiltration

The colonic MPO activity represents another index of inflammation. Colons were processed for MPO activity. TNBS administration resulted in increased MPO activity, indicative of neutrophil infiltration. The increase was significantly higher in mice receiving both TNBS and misoprostol (Fig. 3C). This correlates with the increase in IL-17, whose major function is the attraction and activation of neutrophils.

Misoprostol promotes transmural inflammatory cell infiltration

The colons of mice that received TNBS plus miso exhibited intense necrosis and significantly higher levels of inflammation compared with the TNBS-treated mice (data not shown). Cross-sections of distal colons were stained with H&E. Colons from mice treated with ethanol or with ethanol plus misoprostol show no inflammatory infiltrates and have an intact anatomy of the colonic wall (Fig. 4I, A and B). In contrast, in colons of TNBS-treated mice we observed inflammatory infiltrates and slight tissue disruptions (Fig. 4IC). The pathology is less severe than reported by others because the tissues were harvested 36 h after TNBS administration and not at the time of the maximum clinical score. In contrast, colons from mice given TNBS plus misoprostol showed transmural inflammation, marked increases in the thickness of the muscular layer, epithelial cell loss, pronounced depletion of mucin-producing goblet cells, patchy ulceration, disseminated fibrosis, and focal crypt loss (Fig. 4ID).


Figure 4
View larger version (72K):
[in this window]
[in a new window]

  		FIGURE 4. Histology and immunohistochemistry analyses. I, Colon cryosections were prepared (36 h) and stained with H & E. (A) EtOH, (B) EtOH plus misoprostol, (C) TNBS, and (D) TNBS plus misoprostol. Arrows indicates inflammatory infiltrates. II, Identification of IL-17+ cells. Colon cryosections from EtOH + miso (A, D, and G), TNBS (B, E, and H), or TNBS + miso (C, F, and I) were stained with FITC-conjugated anti-CD4 Ab (1:100; green) (A–C), and PE-conjugated IL-17 Ab (1:100; red) (D–F). Merged images for double-stained cells (yellow) (G–I).

 
Misoprostol leads to a significant increase in the number of IL-17-producing CD4+ T cells in the intestinal mucosa

Colon sections harvested 36 h post-TNBS treatment were stained with FITC-labeled anti-CD4 Abs (Fig. 4II, upper panels) and PE anti-IL-17 Abs (middle panels). Double-stained cells (CD4+IL-17+) are shown in the lower panels. There are no identifiable IL-17 producing CD4+ T cells in mice treated with ethanol plus misoprostol (Fig. 4II, A, D, and G). Thirty-six hours after TNBS administration, there are moderate numbers of CD4+ cells (Fig. 4IIB), of IL-17-producing cells (Fig. 4IIE), and of double-positive cells (Fig. 4IIH). In contrast, following treatment with TNBS and misoprostol, there is a significant influx of CD4+ cells (Fig. 4IIC), high numbers of IL-17-positive cells (Fig. 4IIF), and high numbers of double-positive CD4+IL-17+ cells (Fig. 4III).

Cyclooxygenase (Cox) inhibitors exacerbate TNBS-induced colitis without inducing significant increases in IL-17 and p19 expression

We hypothesized that high levels of PGE2 generated locally in autoimmune/inflammatory conditions affect the IL-12/IL-23 balance in resident DC, promoting a preferential IL-17 T cell response. The data presented above show exacerbation of clinical colitis and increased p19/IL-23 and IL-17 levels upon administration of misoprostol in the TNBS model.

However, several reports indicate that Cox inhibitors, which block endogenous PG release, have a detrimental effect in patients with CrD or UC and in IBD animal models (43, 44, 45, 46, 47). Therefore, we sought to determine whether blockade of endogenous PG synthesis by using Cox inhibitors affects TNBS-induced colitis and whether this is associated with increased IL-23/IL-17 expression. Indomethacin (a nonselective Cox inhibitor) has been used previously used to block PGE2 synthesis in various models, including TNBS-induced colitis (45, 48, 49). Mice were injected i.p. with indomethacin 30 min before TNBS administration, followed by indomethacin inoculations every 12 h on days 1 and 2. Control mice received only EtOH, or EtOH plus indomethacin (no TNBS). Similar to misoprostol, indomethacin exacerbated TNBS-colitis, with 80% of animals succumbing by day 3, and no survivor by day 7 (Fig. 5A). Macroscopic analysis of colons obtained 48 h after indomethacin administration showed hyperemia with prominent hemorrhagic areas (Fig. 5B).


Figure 5
View larger version (26K):
[in this window]
[in a new window]

  		FIGURE 5. Effects of indomethacin in TNBS-induced colitis. BALB/c mice (n = 9) were treated with 50% EtOH, EtOH and indomethacin, TNBS, and TNBS and indomethacin. Groups receiving indomethacin (Indo) were injected with indomethacin (1.5 mg/kg) 30 min before TNBS or EtOH administration, followed by twice daily indomethacin inoculations on days 1 and 2. A, Left panel, Body weight loss on days 3 and 8 after TNBS administration. Numbers in parentheses represent numbers of surviving mice in each group on days 3 and 8, respectively. Right panel, Survival curves. Weight loss was monitored in a blinded fashion. The Kaplan-Meier log-rank test was used for the survival curves. *, p < 0.05 (comparison between indomethacin + TNBS and TNBS treatment). B, Colons harvested 48 h after TNBS administration. C, Expression of p19 and IL-17 in colonic tissue (48 h) and in MLN cells restimulated ex vivo with anti-CD3 and anti-CD28 mAbs for 24 h was analyzed individually by real-time RT-PCR. One representative experiment of two is shown.

 
However, although both indomethacin- and misoprostol-treated mice developed severe TNBS colitis, the cytokine profile turned out to be quite different in the two groups. In contrast to misoprostol, which induces high expression of both p19 and IL-17, indomethacin did not increase IL-17 above TNBS levels in lymph node cells, and resulted in relatively low expression of both p19 and IL-17 in colon (Fig. 5C). These results suggest that high levels of PGE2/misoprostol exacerbate colitis through an inflammatory process involving the IL-23->IL-17 axis, whereas Cox inhibitors which prevent both constitutive and inflammatory endogenous PG synthesis exacerbate colitis through mechanisms unrelated to IL-23/IL-17-induced inflammation.


   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
Disclosures
 References
 
The newly identified CD4+ Th17 cells emerged recently as central players in inflammatory/autoimmune conditions such as rheumatoid arthritis, experimental autoimmune encephalomyelitis, and IBDs (19, 20, 33, 50, 51, 52, 53). Although Th17 differentiation depends primarily on TGFbeta and IL-6, IL-23 is essential for the Th17 expansion and survival (24, 25, 27). In contrast to IL-23, the two other members of the IL-12 family, IL-12p70 and IL-27, suppress the development of Th17 effectors (33, 35, 54). We showed previously that PGE2 induces IL-23, but not IL-12p70, secretion from DC and inhibits IL-12p70 release from LPS-stimulated DC (36). Based on these results, we hypothesized that PGE2 plays a proinflammatory role in inflammatory/autoimmune diseases through the DC-derived IL-23->Th17 axis. In the present study, we show that PGE2 promotes IL-23 and inhibits IL-12/IL-27 expression in LPS-stimulated DC and that PGE2 analogs exacerbate clinical symptoms/histopathology in the TNBS colitis model. In agreement with the proposed effect of PGE2, we found that administration of PGE2 analogs results in an increase in IL-23 and IL-17 and a decrease in IL-12p35 expression in colon and MLN and a substantial increase in the number of colonic CD4+IL-17+ T cells.

Although CrD was previously considered a Th1-type disease (6, 7, 8), there is convincing new evidence that IL-17 plays a central role in CrD. CrD patients have increased levels of IL-17 in serum and intestinal mucosa (16, 17, 18), and treatment with anti-p40 Abs, leading to IL-17 decrease, is beneficial (17). In colitis models, IL-17 was identified in colon and administration of anti-IL-17 Abs suppressed intestinal inflammation (19). TNBS-induced colitis is an established IBD model based on similarities with human CrD (37). Although TNBS colitis is considered a Th1-type disease, the fact that mice expressing high levels of IFN-{gamma} but lacking IL-17R are less susceptible to disease indicates that IL-17, and not IFN-{gamma}, plays the central role (20).

We found that PGE2 inhibits IL-12/IL-27 and promotes IL-23 expression in DC, and that the in vivo administration of PGE analogs results in higher expression of IL-23 and IL-17, and in the accumulation of Th17 cells in the inflamed intestine. This is in agreement with the recently established role of the IL-12 cytokine family in CD4+ T cell differentiation. IL-12 and IL-27 are strong promoters of Th1 differentiation (28, 29, 30, 31, 32), and directly suppress Th17 development (33, 34, 35), possibly through the induction of Tbet which prevents and destabilizes the Th17 phenotype (55, 56). In contrast, IL-23 is required to stabilize the Th17 phenotype (14, 19, 24, 25, 26, 27).

Although the role of IL-23 in IBD has been questioned recently (57), an overwhelming number of clinical and experimental observations support IL-23->Th17 involvement in IBD. Increases in p19 expression were reported in intestinal mucosal biopsies from CrD and UC patients, but not in non-IBD colitis patients (58, 59), and a nonsynonymous variant of the IL-23R has been reported as one of the three markers associated with childhood onset of IBD (the other two markers being located in the NOD2/CARD15 gene) (60). In a model of spontaneous colitis, p19-deficient mice were shown to be disease resistant, and IL-23 administration together with transfer of CD4+CD45RBhigh T cells into Rag-1-deficient mice significantly accelerated disease onset (19). Increased p19 expression was localized to lamina propria CD11c+ F4/80–CD68– cells (61), pointing to DC as the major IL-23 source. In addition, in models of bacterially induced colitis, IL-23 was shown to play a key role in both innate and T cell-mediated intestinal inflammation (62, 63).

The role of IL-12 and IL-23 were addressed recently in a colitis model based strictly on innate immune responses (5). In this model, IL-23 p19 secreted from intestinal DC acted locally, resulting in intestinal inflammation, whereas IL-12 acted systemically inducing serum proinflammatory cytokines and wasting disease. In our hands, exacerbation of TNBS colitis induced by PGE analogs correlates with increased IL-23 and reduced IL-12 expression, with a significant accumulation of Th17 cells, and with the up-regulation of proinflammatory cytokines in the inflamed intestinal tissue. This is in agreement with the proposed local effect of IL-23->IL-17 leading to intestinal inflammation.

PGE2, released at high levels during inflammation, plays mostly a proinflammatory role in vivo. In IBD, however, PGE2 appears to have a dual effect. On one hand, as demonstrated by our data, high levels of PGE2 analogs exacerbate clinical colitis, presumably through the induction of the IL-23->IL-17 proinflammatory axis. On the other hand, constitutive PGE2 production in the intestine appears to have a protective effect on the integrity of the epithelial intestinal wall, presumably through the enhancement of epithelium survival and regeneration (45, 64, 65). This is evident primarily in models of colitis where the central event is damage to the epithelial layer, such as dextran sodium sulfate-induced colitis, and in experimental models using Cox1/2- or EP4-deficient mice, where PGE2 synthesis or signaling of constitutive PGE2 through the EP4 receptor are prevented (65, 66, 67). In agreement with previously reported deleterious effect of nonsteroidal anti-inflammatory drugs or Cox inhibitors in IBD, we found that use of the nonselective Cox inhibitor indomethacin had as drastic an effect in TNBS colitis as the administration of high doses of misoprostol or PGE1-OH. There is however a significant mechanistic difference, with indomethacin having marginal effects on IL-23/IL-17 expression compared with misoprostol. This suggests that indomethacin, and possibly other nonsteroidal anti-inflammatory drugs and Cox inhibitors, act by preventing the beneficial effect of PGE2 and possibly other PG on the repair of the intestinal epithelial cell wall.

Based on our results, we hypothesize that although endogenous low levels of PGE2 are required for intestinal epithelial cell survival and regeneration, high PGE2 levels released locally in inflammatory conditions, such as in established colitis, contribute to the perpetuation of inflammation, exacerbating the disease process through the preferential induction of DC and possibly macrophage-derived IL-23 and subsequent expansion and survival of pathogenic autoimmune Th17 cells. We propose that high levels of PGE2 play a proinflammatory role in IBD by switching the cytokine profile of lamina propria DC (LPDC) (Fig. 6). In their abnormal response to commensal gut flora, LPDC express and secrete all three members of the IL-12 family, i.e., IL-12p70, IL-23, and IL-27. Incoming CD4+ T cells differentiate primarily into Th1 effectors in the presence of IL-12p70 and IL-27. Th17 are also generated due to the presence of TGFbeta, released primarily by colonic subepithelial myofibroblasts (68, 69), and of IL-6 produced by LPDC and activated macrophages. However, following IL-12/IL-27-induced Tbet expression in differentiating T cells, the Th17 phenotype is rapidly destabilized (55). The situation changes in the presence of PGE2. PGE2, generated at high levels during inflammation in IBD, acts on LPDC inhibiting IL-12/IL-27, and up-regulating IL-23 production. This leads to a reduction in Th1 cells and to the stabilization of the Th17 phenotype. In addition to activated T cells, IL-23 also acts on macrophages and DC expressing IL-23R. This leads to the release of IL-1beta and IL-6, required for Th17 differentiation and stabilization. Therefore, in the presence of PGE2 there is preferential production of IL-17 from Th17 effectors, leading to neutrophil accumulation, and activation of fibroblasts, epithelial cells, macrophages to release proinflammatory cytokines and chemokines, as well as metalloproteinases (70). In addition, IL-17 synergizes with LPS to induce Cox2 expression in colonic subepithelial myofibroblasts (71), maintaining a proinflammatory circuit. In conclusion, our model proposes that high levels of PGE2 act as endogenous regulators of the IL-12/IL-27 vs IL-23 balance, promoting the inflammatory IL-23->IL-17 axis, and resulting in locally amplified and sustained inflammation.


Figure 6
View larger version (41K):
[in this window]
[in a new window]

  		FIGURE 6. Model for the PGE2 exacerbation of IBD through the IL-23->IL-17 axis. Aberrant immune responses against commensal gut flora induce IL-12, IL-27, and IL-23 release from LPDC. IL-12/IL-27 promote Th1 and inhibit Th17 differentiation (presumably through Tbet expression). PGE2, released from myofibroblasts, inhibits L-12 and IL-27 release and promotes IL-23 production. IL-23 acts on differentiating Th17 cells, stabilizing their phenotype, and on IL-23R-expressing LPDC and macrophages (M{phi}) inducing IL-6 and IL-1beta release. The combination of IL-23, IL-6, and IL-1beta strongly supports Th17 differentiation and function. IL-17 released from Th17 cells attracts and activates neutrophils, acts on various cells inducing cytokine/chemokine expression and metalloproteinase (MMP) release, and further promotes PGE2 release from myofibroblasts.

 
The fact that high doses of exogenous PGE2 analogs exacerbate clinical colitis in the TNBS model might be relevant to the use of misoprostol to prevent ulcers in patients who take arthritis medication. The side effects listed for misoprostol include a variety of gastrointestinal tract problems, which could very well develop from the unwanted proinflammatory effect mediated by the IL-23->IL-17 axis.


   Disclosures
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
References
 
The authors have no financial conflict of interest.


   Footnotes
 
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

1 This work was supported by National Institutes of Health/National Institute of Allergy and Infectious Diseases Grant RO1AI052306. Back

2 Address correspondence and reprint requests to Dr. Doina Ganea, Department of Physiology, Temple University School of Medicine, 3420 North Broad Street, Philadelphia, PA 19140. E-mail address: doina.ganea{at}temple.edu Back

3 Abbreviations used in this paper: IBD, inflammatory bowel disease; CrD, Crohn’s disease; UC, ulcerative colitis; TNBS, 2,4,6-trinitrobenzene sulfonic acid; DC, dendritic cell; rm, recombinant murine; MPO, myeloperoxidase; MLN, mesenteric lymph node; MPO, myeloperoxidase; LPDC, lamina propria DC; Cox, cyclooxygenase. Back

Received for publication October 30, 2006. Accepted for publication March 30, 2007.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 

  1. Bouma, G., W. Strober. 2003. The immunological and genetic basis of inflammatory bowel disease. Nat. Rev. Immunol. 3: 521-533. [Medline]
  2. Shanahan, F.. 2002. Crohn’s disease. Lancet 359: 62-69. [Medline]
  3. Neurath, M. F., S. Finotto, L. H. Glimcher. 2002. The role of Th1/Th2 polarization in mucosal immunity. Nat. Med. 8: 567-573. [Medline]
  4. Rakoff-Nahoum, S., L. Hao, R. Medzhitov. 2006. Role of Toll-like receptors in spontaneous commensal-dependent colitis. Immunity 25: 319-329. [Medline]
  5. Uhlig, H. H., B. S. McKenzie, S. Hue, C. Thompson, B. Joyce-Shaikh, R. Stepankova, N. Robinson, S. Buonocore, H. Tlaskalova-Hogenova, D. J. Cua, F. Powrie. 2006. Differential activity of IL-12 and IL-23 in mucosal and systemic innate immune pathology. Immunity 25: 309-318. [Medline]
  6. Cobrin, G. M., M. T. Abreu. 2005. Defects in mucosal immunity leading to Crohn’s disease. Immunol. Rev. 206: 277-295. [Medline]
  7. Hanauer, S. B.. 2006. Inflammatory bowel disease: epidemiology, pathogenesis, and therapeutic opportunities. Inflamm. Bowel Dis. 12: (Suppl. 1):S3-S9. [Medline]
  8. Monteleone, G., D. Fina, R. Caruso, F. Pallone. 2006. New mediators of immunity and inflammation in inflammatory bowel disease. Curr. Opin. Gastroenterol. 22: 361-364. [Medline]
  9. Alber, G., S. Al-Robaiy, M. Kleinschek, J. Knauer, P. Krumbholz, J. Richter, S. Schoeneberger, N. Schuetze, S. Schulz, K. Toepfer, et al 2006. Induction of immunity and inflammation by interleukin-12 family members. Ernst. Schering Res. Found. Workshop 56: 107-127. [Medline]
  10. Bowman, E. P., A. A. Chackerian, D. J. Cua. 2006. Rationale and safety of anti-interleukin-23 and anti-interleukin-17A therapy. Curr. Opin. Infect. Dis. 19: 245-252. [Medline]
  11. Chen, Y., C. L. Langrish, B. McKenzie, B. Joyce-Shaikh, J. S. Stumhofer, T. McClanahan, W. Blumenschein, T. Churakovsa, J. Low, L. Presta, et al 2006. Anti-IL-23 therapy inhibits multiple inflammatory pathways and ameliorates autoimmune encephalomyelitis. J. Clin. Invest. 116: 1317-1326. [Medline]
  12. Cua, D. J., J. Sherlock, Y. Chen, C. A. Murphy, B. Joyce, B. Seymour, L. Lucian, W. To, S. Kwan, T. Churakova, et al 2003. Interleukin-23 rather than interleukin-12 is the critical cytokine for autoimmune inflammation of the brain. Nature 421: 744-748. [Medline]
  13. Harrington, L. E., P. R. Mangan, C. T. Weaver. 2006. Expanding the effector CD4 T-cell repertoire: the Th17 lineage. Curr. Opin. Immunol. 18: 349-356. [Medline]
  14. Langrish, C. L., Y. Chen, W. M. Blumenschein, J. Mattson, B. Basham, J. D. Sedgwick, T. McClanahan, R. A. Kastelein, D. J. Cua. 2005. IL-23 drives a pathogenic T cell population that induces autoimmune inflammation. J. Exp. Med. 201: 233-240. [Abstract/Free Full Text]
  15. Murphy, C. A., C. L. Langrish, Y. Chen, W. Blumenschein, T. McClanahan, R. A. Kastelein, J. D. Sedgwick, D. J. Cua. 2003. Divergent pro- and antiinflammatory roles for IL-23 and IL-12 in joint autoimmune inflammation. J. Exp. Med. 198: 1951-1957. [Abstract/Free Full Text]
  16. Fujino, S., A. Andoh, S. Bamba, A. Ogawa, K. Hata, Y. Araki, T. Bamba, Y. Fujiyama. 2003. Increased expression of interleukin 17 in inflammatory bowel disease. Gut 52: 65-70. [Abstract/Free Full Text]
  17. Fuss, I. J., C. Becker, Z. Yang, C. Groden, R. L. Hornung, F. Heller, M. F. Neurath, W. Strober, P. J. Mannon. 2006. Both IL-12p70 and IL-23 are synthesized during active Crohn’s disease and are down-regulated by treatment with anti-IL-12 p40 monoclonal antibody. Inflamm. Bowel Dis. 12: 9-15. [Medline]
  18. Nielsen, O. H., I. Kirman, N. Rudiger, J. Hendel, B. Vainer. 2003. Upregulation of interleukin-12 and -17 in active inflammatory bowel disease. Scand. J. Gastroenterol. 38: 180-185. [Medline]
  19. Yen, D., J. Cheung, H. Scheerens, F. Poulet, T. McClanahan, B. McKenzie, M. A. Kleinschek, A. Owyang, J. Mattson, W. Blumenschein, et al 2006. IL-23 is essential for T cell-mediated colitis and promotes inflammation via IL-17 and IL-6. J. Clin. Invest. 116: 1310-1316. [Medline]
  20. Zhang, Z., M. Zheng, J. Bindas, P. Schwarzenberger, J. K. Kolls. 2006. Critical role of IL-17 receptor signaling in acute TNBS-induced colitis. Inflamm. Bowel Dis. 12: 382-388. [Medline]
  21. Beadling, C., M. K. Slifka. 2006. Regulation of innate and adaptive immune responses by the related cytokines IL-12, IL-23, and IL-27. Arch. Immunol. Ther. Exp. 54: 15-24.
  22. Hunter, C. A.. 2005. New IL-12-family members: IL-23 and IL-27, cytokines with divergent functions. Nat. Rev. Immunol. 5: 521-531. [Medline]
  23. Kang, B. Y., T. S. Kim. 2006. Targeting cytokines of the interleukin-12 family in autoimmunity. Curr. Med. Chem. 13: 1149-1156. [Medline]
  24. Bettelli, E., Y. Carrier, W. Gao, T. Korn, T. B. Strom, M. Oukka, H. L. Weiner, V. K. Kuchroo. 2006. Reciprocal developmental pathways for the generation of pathogenic effector TH17 and regulatory T cells. Nature 441: 235-238. [Medline]
  25. Mangan, P. R., L. E. Harrington, D. B. O’Quinn, W. S. Helms, D. C. Bullard, C. O. Elson, R. D. Hatton, S. M. Wahl, T. R. Schoeb, C. T. Weaver. 2006. Transforming growth factor-beta induces development of the Th17 lineage. Nature 441: 231-234. [Medline]
  26. McKenzie, B. S., R. A. Kastelein, D. J. Cua. 2006. Understanding the IL-23-IL-17 immune pathway. Trends Immunol. 27: 17-23. [Medline]
  27. Veldhoen, M., R. J. Hocking, C. J. Atkins, R. M. Locksley, B. Stockinger. 2006. TGFbeta in the context of an inflammatory cytokine milieu supports de novo differentiation of IL-17-producing T cells. Immunity 24: 179-189. [Medline]
  28. Pflanz, S., J. C. Timans, J. Cheung, R. Rosales, H. Kanzler, J. Gilbert, L. Hibbert, T. Churakova, M. Travis, E. Vaisberg, et al 2002. IL-27, a heterodimeric cytokine composed of EBI3 and p28 protein, induces proliferation of naive CD4+ T cells. Immunity 16: 779-790. [Medline]
  29. Takeda, A., S. Hamano, A. Yamanaka, T. Hanada, T. Ishibashi, T. W. Mak, A. Yoshimura, H. Yoshida. 2003. Cutting edge: role of IL-27/WSX-1 signaling for induction of T-bet through activation of STAT1 during initial Th1 commitment. J. Immunol. 170: 4886-4890. [Abstract/Free Full Text]
  30. Trinchieri, G.. 2003. Interleukin-12 and the regulation of innate resistance and adaptive immunity. Nat. Rev. Immunol. 3: 133-146. [Medline]
  31. Trinchieri, G., S. Pflanz, R. A. Kastelein. 2003. The IL-12 family of heterodimeric cytokines: new players in the regulation of T cell responses. Immunity 19: 641-644. [Medline]
  32. Villarino, A. V., J. Larkin, 3rd, C. J. Saris, A. J. Caton, S. Lucas, T. Wong, F. J. de Sauvage, C. A. Hunter. 2005. Positive and negative regulation of the IL-27 receptor during lymphoid cell activation. J. Immunol. 174: 7684-7691. [Abstract/Free Full Text]
  33. Batten, M., J. Li, S. Yi, N. M. Kljavin, D. M. Danilenko, S. Lucas, J. Lee, F. J. de Sauvage, N. Ghilardi. 2006. Interleukin 27 limits autoimmune encephalomyelitis by suppressing the development of interleukin 17-producing T cells. Nat. Immunol. 7: 929-936. [Medline]
  34. Hoeve, M. A., N. D. Savage, T. de Boer, D. M. Langenberg, R. de Waal Malefyt, T. H. Ottenhoff, F. A. Verreck. 2006. Divergent effects of IL-12 and IL-23 on the production of IL-17 by human T cells. Eur. J. Immunol. 36: 661-670. [Medline]
  35. Stumhofer, J. S., A. Laurence, E. H. Wilson, E. Huang, C. M. Tato, L. M. Johnson, A. V. Villarino, Q. Huang, A. Yoshimura, D. Sehy, et al 2006. Interleukin 27 negatively regulates the development of interleukin 17-producing T helper cells during chronic inflammation of the central nervous system. Nat. Immunol. 7: 937-945. [Medline]
  36. Sheibanie, A. F., I. Tadmori, H. Jing, E. Vassiliou, D. Ganea. 2004. Prostaglandin E2 induces IL-23 production in bone marrow-derived dendritic cells. FASEB J. 18: 1318-1320. [Abstract/Free Full Text]
  37. Neurath, M., I. Fuss, W. Strober. 2000. TNBS-colitis. Int. Rev. Immunol. 19: 51-62. [Medline]
  38. Jing, H., E. Vassiliou, D. Ganea. 2003. Prostaglandin E2 inhibits production of the inflammatory chemokines CCL3 and CCL4 in dendritic cells. J. Leukocyte Biol. 74: 868-879. [Abstract/Free Full Text]
  39. Chin, K. W., K. E. Barrett. 1994. Mast cells are not essential to inflammation in murine model of colitis. Dig. Dis. Sci. 39: 513-525. [Medline]
  40. Gonzalez-Rey, E., N. Varela, A. F. Sheibanie, A. Chorny, D. Ganea, M. Delgado. 2006. Cortistatin, an antiinflammatory peptide with therapeutic action in inflammatory bowel disease. Proc. Natl. Acad. Sci. USA 103: 4228-4233. [Abstract/Free Full Text]
  41. Vassiliou, E., H. Jing, D. Ganea. 2003. Prostaglandin E2 inhibits TNF production in murine bone marrow-derived dendritic cells. Cell. Immunol. 223: 120-132. [Medline]
  42. Kiriyama, M., F. Ushikubi, T. Kobayashi, M. Hirata, Y. Sugimoto, S. Narumiya. 1997. Ligand binding specificities of the eight types and subtypes of the mouse prostanoid receptors expressed in Chinese hamster ovary cells. Br. J. Pharmacol. 122: 217-224. [Medline]
  43. Meyer, A. M., N. N. Ramzan, R. I. Heigh, J. A. Leighton. 2006. Relapse of inflammatory bowel disease associated with use of nonsteroidal anti-inflammatory drugs. Dig. Dis. Sci. 51: 168-172. [Medline]
  44. Shafiq, N., S. Malhotra, P. Pandhi, R. Nada. 2005. Comparative gastrointestinal toxicity of selective cyclooxygenase (COX-2) inhibitors. Indian J. Exp. Biol. 43: 614-619. [Medline]
  45. Tessner, T. G., S. M. Cohn, S. Schloemann, W. F. Stenson. 1998. Prostaglandins prevent decreased epithelial cell proliferation associated with dextran sodium sulfate injury in mice. Gastroenterology 115: 874-882. [Medline]
  46. Thiefin, G., L. Beaugerie. 2005. Toxic effects of nonsteroidal antiinflammatory drugs on the small bowel, colon, and rectum. Joint Bone Spine 72: 286-294. [Medline]
  47. Zhang, L., Y. M. Lu, X. Y. Dong. 2004. Effects and mechanism of the selective COX-2 inhibitor, celecoxib, on rat colitis induced by trinitrobenzene sulfonic acid. Chin. J. Dig. Dis. 5: 110-114. [Medline]
  48. Ballinger, M. N., D. M. Aronoff, T. R. McMillan, K. R. Cooke, K. Olkiewicz, G. B. Toews, M. Peters-Golden, B. B. Moore. 2006. Critical role of prostaglandin E2 overproduction in impaired pulmonary host response following bone marrow transplantation. J. Immunol. 177: 5499-5508. [Abstract/Free Full Text]
  49. Kankuri, E., K. Vaali, R. Korpela, I. Paakkari, H. Vapaatalo, E. Moilanen. 2001. Effects of a COX-2 preferential agent nimesulide on TNBS-induced acute inflammation in the gut. Inflammation 25: 301-310. [Medline]
  50. Komiyama, Y., S. Nakae, T. Matsuki, A. Nambu, H. Ishigame, S. Kakuta, K. Sudo, Y. Iwakura. 2006. IL-17 plays an important role in the development of experimental autoimmune encephalomyelitis. J. Immunol. 177: 566-573. [Abstract/Free Full Text]
  51. Lubberts, E., M. I. Koenders, W. B. van den Berg. 2005. The role of T-cell interleukin-17 in conducting destructive arthritis: lessons from animal models. Arthritis Res. Ther. 7: 29-37. [Medline]
  52. Park, H., Z. Li, X. O. Yang, S. H. Chang, R. Nurieva, Y. H. Wang, Y. Wang, L. Hood, Z. Zhu, Q. Tian, C. Dong. 2005. A distinct lineage of CD4 T cells regulates tissue inflammation by producing interleukin 17. Nat. Immunol. 6: 1133-1141. [Medline]
  53. Sutton, C., C. Brereton, B. Keogh, K. H. Mills, E. C. Lavelle. 2006. A crucial role for interleukin (IL)-1 in the induction of IL-17-producing T cells that mediate autoimmune encephalomyelitis. J. Exp. Med. 203: 1685-1691. [Abstract/Free Full Text]
  54. Cruz, A., S. A. Khader, E. Torrado, A. Fraga, J. E. Pearl, J. Pedrosa, A. M. Cooper, A. G. Castro. 2006. Cutting edge: IFN-{gamma} regulates the induction and expansion of IL-17-producing CD4 T cells during mycobacterial infection. J. Immunol. 177: 1416-1420. [Abstract/Free Full Text]
  55. Mathur, A. N., H. C. Chang, D. G. Zisoulis, R. Kapur, M. L. Belladonna, G. S. Kansas, M. H. Kaplan. 2006. T-bet is a critical determinant in the instability of the IL-17-secreting T-helper phenotype. Blood 108: 1595-1601. [Abstract/Free Full Text]
  56. Rangachari, M., N. Mauermann, R. R. Marty, S. Dirnhofer, M. O. Kurrer, V. Komnenovic, J. M. Penninger, U. Eriksson. 2006. T-bet negatively regulates autoimmune myocarditis by suppressing local production of interleukin 17. J. Exp. Med. 203: 2009-2019. [Abstract/Free Full Text]
  57. Becker, C., H. Dornhoff, C. Neufert, M. C. Fantini, S. Wirtz, S. Huebner, A. Nikolaev, H. A. Lehr, A. J. Murphy, D. M. Valenzuela, et al 2006. Cutting edge: IL-23 cross-regulates IL-12 production in T cell-dependent experimental colitis. J. Immunol. 177: 2760-2764. [Abstract/Free Full Text]
  58. Schmidt, C., T. Giese, B. Ludwig, I. Mueller-Molaian, T. Marth, S. Zeuzem, S. C. Meuer, A. Stallmach. 2005. Expression of interleukin-12-related cytokine transcripts in inflammatory bowel disease: elevated interleukin-23p19 and interleukin-27p28 in Crohn’s disease but not in ulcerative colitis. Inflamm. Bowel Dis. 11: 16-23. [Medline]
  59. Stallmach, A., T. Giese, C. Schmidt, B. Ludwig, I. Mueller-Molaian, S. C. Meuer. 2004. Cytokine/chemokine transcript profiles reflect mucosal inflammation in Crohn’s disease. Int. J. Colorectal Dis. 19: 308-315. [Medline]
  60. Van Limbergen, J. E., R. K. Russell, E. R. Nimmo, H. E. Drummond, L. Smith, N. H. Anderson, G. Davies, P. M. Gillett, P. McGrogan, K. Hassan, et al. 2007. IL23R Arg381Gln is associated with childhood onset inflammatory bowel disease in Scotland. Gut. In press.
  61. Krajina, T., F. Leithauser, P. Moller, Z. Trobonjaca, J. Reimann. 2003. Colonic lamina propria dendritic cells in mice with CD4+ T cell-induced colitis. Eur. J. Immunol. 33: 1073-1083. [Medline]
  62. Hue, S., P. Ahern, S. Buonocore, M. C. Kullberg, D. J. Cua, B. S. McKenzie, F. Powrie, K. J. Maloy. 2006. Interleukin-23 drives innate and T cell-mediated intestinal inflammation. J. Exp. Med. 203: 2473-2483. [Abstract/Free Full Text]
  63. Kullberg, M. C., D. Jankovic, C. G. Feng, S. Hue, P. L. Gorelick, B. S. McKenzie, D. J. Cua, F. Powrie, A. W. Cheever, K. J. Maloy, A. Sher. 2006. IL-23 plays a key role in Helicobacter hepaticus-induced T cell-dependent colitis. J. Exp. Med. 203: 2485-2494. [Abstract/Free Full Text]
  64. Cohn, S. M., S. Schloemann, T. Tessner, K. Seibert, W. F. Stenson. 1997. Crypt stem cell survival in the mouse intestinal epithelium is regulated by prostaglandins synthesized through cyclooxygenase-1. J. Clin. Invest. 99: 1367-1379. [Medline]
  65. Jiang, G. L., A. Nieves, W. B. Im, D. W. Old, D. T. Dinh, L. Wheeler. 2007. The prevention of colitis by E prostanoid receptor 4 agonist through enhancement of epithelium survival and regeneration. J. Pharmacol. Exp. Ther. 320: 22-28. [Abstract/Free Full Text]
  66. Kabashima, K., T. Saji, T. Murata, M. Nagamachi, T. Matsuoka, E. Segi, K. Tsuboi, Y. Sugimoto, T. Kobayashi, Y. Miyachi, et al 2002. The prostaglandin receptor EP4 suppresses colitis, mucosal damage and CD4 cell activation in the gut. J. Clin. Invest. 109: 883-893. [Medline]
  67. Morteau, O., S. G. Morham, R. Sellon, L. A. Dieleman, R. Langenbach, O. Smithies, R. B. Sartor. 2000. Impaired mucosal defense to acute colonic injury in mice lacking cyclooxygenase-1 or cyclooxygenase-2. J. Clin. Invest. 105: 469-478. [Medline]
  68. Beltinger, J., B. C. McKaig, S. Makh, W. A. Stack, C. J. Hawkey, Y. R. Mahida. 1999. Human colonic subepithelial myofibroblasts modulate transepithelial resistance and secretory response. Am. J. Physiol. 277: C271-C279. [Medline]
  69. McKaig, B. C., K. Hughes, P. J. Tighe, Y. R. Mahida. 2002. Differential expression of TGF-beta isoforms by normal and inflammatory bowel disease intestinal myofibroblasts. Am. J. Physiol. 282: C172-C182.
  70. Iwakura, Y., H. Ishigame. 2006. The IL-23/IL-17 axis in inflammation. J. Clin. Invest. 116: 1218-1222. [Medline]
  71. Zhang, Z., A. Andoh, O. Inatomi, S. Bamba, A. Takayanagi, N. Shimizu, Y. Fujiyama. 2005. Interleukin-17 and lipopolysaccharides synergistically induce cyclooxygenase-2 expression in human intestinal myofibroblasts. J. Gastroenterol. Hepatol. 20: 619-627. [Medline]



This article has been cited by other articles:


Home page
 Int ImmunolHome page
H. Shiraishi, H. Yoshida, K. Saeki, Y. Miura, S. Watanabe, T. Ishizaki, M. Hashimoto, G. Takaesu, T. Kobayashi, and A. Yoshimura
Prostaglandin E2 is a major soluble factor produced by stromal cells for preventing inflammatory cytokine production from dendritic cells
Int. Immunol., July 17, 2008; (2008) dxn078v1.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
T. Khayrullina, J.-H. Yen, H. Jing, and D. Ganea
In Vitro Differentiation of Dendritic Cells in the Presence of Prostaglandin E2 Alters the IL-12/IL-23 Balance and Promotes Differentiation of Th17 Cells
J. Immunol., July 1, 2008; 181(1): 721 - 735.
[Abstract] [Full Text] [PDF]

Home page
 Infect. Immun.Home page
M. D. Woolard, L. L. Hensley, T. H. Kawula, and J. A. Frelinger
Respiratory Francisella tularensis Live Vaccine Strain Infection Induces Th17 Cells and Prostaglandin E2, Which Inhibits Generation of Gamma Interferon-Positive T Cells
Infect. Immun., June 1, 2008; 76(6): 2651 - 2659.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Sheibanie, A. F.
Right arrow Articles by Ganea, D.
Right arrow Search for Related Content
PubMed