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. 2019 Feb;143(2):631-643.
doi: 10.1016/j.jaci.2018.05.036. Epub 2018 Jun 20.

T Cell-Intrinsic Prostaglandin E 2-EP2/EP4 Signaling Is Critical in Pathogenic T H 17 Cell-Driven Inflammation

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Free PMC article

T Cell-Intrinsic Prostaglandin E 2-EP2/EP4 Signaling Is Critical in Pathogenic T H 17 Cell-Driven Inflammation

Jinju Lee et al. J Allergy Clin Immunol. .
Free PMC article

Abstract

Background: IL-23 is the key cytokine for generation of pathogenic IL-17-producing helper T (TH17) cells, which contribute critically to autoimmune diseases. However, how IL-23 generates pathogenic TH17 cells remains to be elucidated.

Objectives: We sought to examine the involvement, molecular mechanisms, and clinical implications of prostaglandin (PG) E2-EP2/EP4 signaling in induction of IL-23-driven pathogenic TH17 cells.

Methods: The role of PGE2 in induction of pathogenic TH17 cells was investigated in mouse TH17 cells in culture in vitro and in an IL-23-induced psoriasis mouse model in vivo. Clinical relevance of the findings in mice was examined by using gene expression profiling of IL-23 and PGE2-EP2/EP4 signaling in psoriatic skin from patients.

Results: IL-23 induces Ptgs2, encoding COX2 in TH17 cells, and produces PGE2, which acts back on the PGE receptors EP2 and EP4 in these cells and enhances IL-23-induced expression of an IL-23 receptor subunit gene, Il23r, by activating signal transducer and activator of transcription (STAT) 3, cAMP-responsive element binding protein 1, and nuclear factor κ light chain enhancer of activated B cells (NF-κB) through cyclic AMP-protein kinase A signaling. This PGE2 signaling also induces expression of various inflammation-related genes, which possibly function in TH17 cell-mediated pathology. Combined deletion of EP2 and EP4 selectively in T cells suppressed accumulation of IL-17A+ and IL-17A+IFN-γ+ pathogenic Th17 cells and abolished skin inflammation in an IL-23-induced psoriasis mouse model. Analysis of human psoriatic skin biopsy specimens shows positive correlation between PGE2 signaling and the IL-23/TH17 pathway.

Conclusions: T cell-intrinsic EP2/EP4 signaling is critical in IL-23-driven generation of pathogenic TH17 cells and consequent pathogenesis in the skin.

Keywords: IL-23 receptor; Psoriasis; cAMP-responsive element binding protein 1; nuclear factor κ light chain enhancer of activated B cells; pathogenic T(H)17 cells; prostaglandin E receptor EP2; prostaglandin E receptor EP4; prostaglandin E(2); signal transducer and activator of transcription 3.

Figures

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Fig 1
Fig 1
IL-23 mobilizes the endogenous PGE2-EP2/EP4-cAMP-PKA pathway to facilitate TH17 expansion through synergistic Il23r induction. A and B, Expansion of the TH17 population by PGE2 and IL-23. CD4+ T cells were differentiated with TGF-β1 and IL-6 to TH17 cells for 4 days and then stimulated with 100 nmol/L PGE2 in the absence or presence of IL-23 (10 ng/mL) for an additional 3 days. The cells were examined by using fluorescence-activated cell sorting for IL-17A and IFN-γ (Fig 1, A) and by using quantitative RT-PCR for Il17a expression (Fig 1, B). C-E, Effects of PGE2, 100 μM of an agonist selective to each EP subtype, and related compounds on Il23r expression. TH17 cells were incubated with 100 nmol/L PGE2, an agonist selective to each EP subtype, ONO-DI-004 (EP1), ONO-AE1-259 (EP2), ONO-AE-248 (EP3), or ONO-AE1-329 (EP4), 100 μmol/L db-cAMP, 10 μmol/L forskolin (FSK), or 100 μmol/L 3-isobutyl-1-methylxanthine (IBMX) with or without IL-23. Il23r expression (Fig 1, C and D) or IL-17A concentrations in culture supernatants (Fig 1, E) were examined. F and G, Expression of Il23r in TH17 cells stimulated with 100 μmol/L db-cAMP, 300 μmol/L N6-Bnz-cAMP (a PKA agonist), 300 μmol/L 8-pCTP-2′-O-Me-cAMP (an Epac activator; Fig 1, F), or 10 μM H-89 (a PKA inhibitor; Fig 1, G) with or without IL-23. All bars indicate means ± SEMs (n = 3). *P < .05, **P < .01, and ***P < .001.
Fig 2
Fig 2
IL-23 self-amplifies its own signaling through a T cell–intrinsic positive feedback COX2–PGE2–cAMP–IL-23R loop. A, Expression of COX2 mRNA (Ptgs2) in TH17 cells or TH17 cells cultured further in the presence or absence of IL-23 for 3 days, as determined by using quantitative RT-PCR. B, Concentrations of PGE2 in culture supernatants of TH17 cells in the presence or absence of IL-23 and indomethacin determined by means of ELISA. n.d., Not detected. C,Il23r expression in TH17 cells stimulated with PGE2 and IL-23 in the presence or absence of 100 μM indomethacin for 3 days. D,Il23r expression in TH17 cells stimulated with PGE2 and IL-23 in the presence or absence of EP2 (PF-04418948, 300 μM) and/or EP4 (ONO-AE3-208, 100 μM) antagonists for 3 days. All bars indicate means ± SEMs (n = 3). *P < .05, **P < .01, and ***P < .001.
Fig 3
Fig 3
STAT3, CREB1, and NF-κB mediate cAMP- and IL-23–induced Il23r expression in TH17 cells. A, Effect of STAT3 inhibitor VII on Il23r expression in TH17 cells stimulated with db-cAMP, IL-23, or both for 3 days. B, Western blot analysis of phospho-Y705 STAT3 and α-tubulin as a loading control in TH17 cells cultured as described in the Methods section in this article's Online Repository. Representative images from 2 independent experiments are shown. C, Effect of KG-501 on Il23r expression in TH17 cells stimulated by using db-cAMP, IL-23, or both. D, Effects of RNA interference for CREB1 on Il23r expression (left) and Creb1 expression to confirm CREB knockdown efficiency (right). RNA interference, subsequent culture, and stimulation of TH17 cells were performed, as described in the Methods section in this article's Online Repository. E, Western blot analysis of phospho-S536 NF-κB p65 (pp65), phospho-S933 NF-κB p105 (pp105), p65, p105/p50, and α-tubulin in TH17 cells stimulated as described in the Methods section in this article's Online Repository. Representative images from 2 independent experiments are shown. F and G, Effects of p105 KO (Fig 3, F) or 50 μM CTP-NBD (Fig 3, G) on Il23r expression in TH17 cells stimulated with db-cAMP, IL-23, or both for 3 days. All bars indicate means ± SEMs (n = 3 for Fig 3, A, C, F, and G; n = 18 in Fig 3, B). ***P < .001.
Fig 4
Fig 4
Activation of the COX2-PGE2-EP2/EP4-cAMP pathway confers a pathogenic TH17 phenotype. A, Microarray analysis of gene expression profiles in TH17 cells stimulated with db-cAMP, IL-23, or both. Venn diagram analysis of 2-fold upregulated or downregulated genes compared with the vehicle control (Veh) on each stimulus (P < .05, 1-way ANOVA; n = 3; left and right, respectively). B, Heat map analysis of expression of selected genes from each cluster. C, Quantitative RT-PCR analysis of expression of representative genes of TH17 signature and immune activation in response to db-cAMP, IL-23, or db-cAMP and IL-23 in combination. D, Quantitative RT-PCR analysis of expression of a representative inflammation suppressor gene, Il10, in response to db-cAMP, IL-23, or db-cAMP and IL-23 in combination. All bars in Fig 4, C and D, indicate means ± SEMs (n = 3). *P < .05, **P < .01, and ***P < .001.
Fig 5
Fig 5
PGE2-EP2/EP4 signaling in T cells is required for IL-23–driven psoriatic skin inflammation. A-D, Ear swelling (n = 16-17; Fig 5, A), representative hematoxylin and eosin staining of the histologic section of the ear (n = 3-4; Fig 5, B), number of IL-17A+ and IL-17A+IFN-γ+CD4+ T cells of the ear (Fig 5, C), and gene expression of Il17a, Ifng, and Il23r in whole ear tissue (Fig 5, D) of WT or EP2 KO mice subcutaneously injected with IL-23 or PBS into the ear daily. An EP4 antagonist (AS1954813, 100 mg/kg) or vehicle was orally administered twice a day to the indicated mice. Bars in Fig 5, B = 50 μm. Representative quantification results of the cell number in each population from 4 independent fluorescence-activated cell sorting experiments are shown in Fig 5, C (n = 3). Gene expression was indicated as fold change compared with PBS-injected ears in Fig 5, D (n = 3). E and F, EP2flox/floxEP4flox/floxLck-Cre+ mice and control WT Lck-Cre+ mice were subjected to an IL-23–induced psoriasis model, and ear swelling (n = 11 and 7, respectively; Fig 5, E) and numbers of IL-17A+ and IL-17A+IFN-γ+CD4+ T cells in the ear (n = 7 and 3, respectively; Fig 5, F) were analyzed. All bars indicate means ± SEMs. *P < .05, **P < .01, and ***P < .001.
Fig 6
Fig 6
PGE2 signaling positively correlates with the IL-23/TH17 pathway in human psoriatic skin biopsy specimens. A, Expression profiles of genes related to PGE2 signaling and TH17 signature genes in human nonlesional (NL) or lesional (PL) skin biopsy specimens from patients with psoriasis (n = 58) and skin samples from healthy control subjects (HC; n = 64). The z score transformed values of the microarray gene expression data set GSE13355 were used. The TH17 score was generated based on the average expression level of IL23A, IL12B, IL23R, IL17A, IL17F, and IL22 genes. B, Correlations of PTGES, PTGES2, HPGD, and PTGER4 gene expression versus those of the TH17 score. Black, green, and red dots indicate healthy control, nonlesional, and lesional psoriatic biopsy specimens, respectively. C, Expression profiles of genes related to PGE2 synthases and TH17 signature genes in human lesional skin biopsy specimens from patients with moderate-to-severe psoriasis before (Baseline, n = 22) or 12 weeks after treatment with the IL-23–specific mAb guselkumab (n = 8). The z score transformed values of the microarray gene expression data set GSE51440 were used. D, Correlations of gene expression of PTGS2 and PTGES versus that of IL23R. P values were calculated by using nonparametric Wilcoxon-Mann-Whitney tests (Fig 6, A and C) or the nonparametric Spearman correlation test (Fig 6, B and D).

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References

    1. Dong C. TH17 cells in development: an updated view of their molecular identity and genetic programming. Nat Rev Immunol. 2008;8:337–348. - PubMed
    1. Kemper C., Atkinson J.P. T-cell regulation: with complements from innate immunity. Nat Rev Immunol. 2007;7:9–18. - PubMed
    1. Korn T., Bettelli E., Oukka M., Kuchroo V.K. IL-17 and Th17 cells. Annu Rev Immunol. 2009;27:485–517. - PubMed
    1. Zhu J., Paul W.E. Peripheral CD4+ T-cell differentiation regulated by networks of cytokines and transcription factors. Immunol Rev. 2010;238:247–262. - PMC - PubMed
    1. Cua D.J., Tato C.M. Innate IL-17-producing cells: the sentinels of the immune system. Nat Rev Immunol. 2010;10:479–489. - PubMed

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