Neuropeptide signaling through neurokinin-1 and neurokinin-2 receptors augments antigen presentation by human dendritic cells
2015; Elsevier BV; Volume: 136; Issue: 6 Linguagem: Inglês
10.1016/j.jaci.2015.06.050
ISSN1097-6825
AutoresJunya Ohtake, Shun Kaneumi, Mishie Tanino, Takuto Kishikawa, Satoshi Terada, Kentaro Sumida, Kazutaka Masuko, Yosuke Ohno, Toshiyuki Kita, Sadahiro Iwabuchi, Toshiya Shinohara, Yoshinori Tanino, Tamiko Takemura, Shinya Tanaka, Hiroya Kobayashi, Hidemitsu Kitamura,
Tópico(s)Neuropeptides and Animal Physiology
ResumoAsthma, an inflammatory disease of the airways, is associated with various TH and innate immune cells.1Holgate S.T. Innate and adaptive immune responses in asthma.Nat Med. 2012; 18: 673-683Crossref PubMed Scopus (610) Google Scholar Pulmonary disorders with chronic inflammation induced by infections and severe asthma, including childhood asthma and chronic obstructive pulmonary disease in smokers, are major public health issues. Hypersensitivity pneumonitis (HP) is a complex pulmonary disease caused by repeated exposure to allergens and irritants.2Lacasse Y. Girard M. Cormier Y. Recent advances in hypersensitivity pneumonitis.Chest. 2012; 142: 208-217Abstract Full Text Full Text PDF PubMed Scopus (129) Google Scholar The precise mechanisms mediating severe asthma and HP remain to be elucidated. Neurotransmitters including substance P (SP) and neurokinin A (NKA) are widely distributed in both the central and peripheral nervous systems, and their receptors, neurokinin-1 receptor (NK1R) and neurokinin-2 receptor (NK2R), are expressed on immune cells. However, the role of the NKA-NK2R axis in immune responses relative to the SP-NK1R signaling cascade has not been elucidated. Previously, we demonstrated that neuropeptide signaling through NK2R was correlated with airway hyperresponsiveness in mouse with severe asthma models.3Kobayashi M. Ashino S. Shiohama Y. Wakita D. Kitamura H. Nishimura T. IFN-gamma elevates airway hyper-responsiveness via up-regulation of neurokinin A/neurokinin-2 receptor signaling in a severe asthma model.Eur J Immunol. 2012; 42: 393-402Crossref PubMed Scopus (21) Google Scholar Moreover, we found that the NKA-NK2R signaling pathway was involved in the antigen-presenting function of murine dendritic cells (DCs) and that NK2R expression on DCs was enhanced by IFN-γ and LPS stimulation in a signal transducer and activator of transcription 1 (STAT-1)-dependent manner.4Kitamura H. Kobayashi M. Wakita D. Nishimura T. Neuropeptide signaling activates dendritic cell-mediated type 1 immune responses through neurokinin-2 receptor.J Immunol. 2012; 188: 4200-4208Crossref PubMed Scopus (17) Google Scholar In addition, we confirmed that human DCs significantly upregulated gene expression levels of NK2R and tachykinin, precursor 1 (TAC-1), which encoded SP and NKA, after IFN-γ or poly I:C stimulation, suggesting possible roles of neuropeptide signaling in human DCs.4Kitamura H. Kobayashi M. Wakita D. Nishimura T. Neuropeptide signaling activates dendritic cell-mediated type 1 immune responses through neurokinin-2 receptor.J Immunol. 2012; 188: 4200-4208Crossref PubMed Scopus (17) Google Scholar In this study, we investigated the effects of neuropeptide signaling through NK2R as well as NK1R on antigen presentation by human DCs and the subsequent activation of effector TH cells. Human DCs (monocyte-derived dendritic cells [MoDCs]), generated from PBMCs in the presence of GM-CSF plus IL-4, demonstrated increased NK2R and NK1R expression after IFN-β, IFN-γ, LPS, or poly I:C stimulation (Fig 1, A and B). The upregulation by poly I:C stimulation was convincing, whereas effects of IFNs were modest. In this study, we found that poly I:C–stimulated MoDCs also induced IFN-α and IFN-β production (see Fig E1, A, in this article's Online Repository at www.jacionline.org). Upregulation of NK1R and NK2R gene expression by poly I:C was STAT-1 dependent (Fig 1, C). We confirmed that surface expression levels of NK1R and NK2R on MoDCs were significantly enhanced by poly I:C stimulation (Fig 1, D). In addition, we found that both NK2R and NK1R were expressed on CD68- or CD163-positive macrophages and HLA-DR–positive cells including DCs in lung tissues from patients with asthma and acute and chronic HP (Fig 1, E and F; see Fig E2 and Tables E1 and E2 in this article's Online Repository at www.jacionline.org). These findings suggest that STAT1 activation by TH1 cytokines, including IFN-γ and type 1 IFNs induced by Toll-like receptor ligands such as poly I:C, drives the upregulation of NK1R and NK2R on DCs. To further evaluate whether neuropeptide signaling through NK1R and NK2R was related to poly I:C–induced DC maturation, we analyzed the surface expression levels of HLA and costimulatory molecules after treatment with an NK1R or NK2R antagonist. HLA-DR, CD80, CD86, and CD40 expressions, but not HLA-A/B/C and CCR7 expressions, upregulated by poly I:C stimulation through STAT1 activation (see Fig E1, B), were attenuated in the presence of an NK1R or NK2R antagonist (Fig 2, A). In this experiment, we confirmed that these antagonists did not increase 7AAD+ and/or Annexin+ cells after the treatments. We further confirmed that SP (25.2 ± 9.2 pg/mL) and NKA (44.6 ± 27.5 pg/mL) were produced in the culture supernatants of MoDCs (2.5 × 105 cells) for 24 hours and HLA class II expression levels on these neuropeptide-treated MoDCs were suppressed in the presence of these antagonists (see Fig E3 in this article's Online Repository at www.jacionline.org). Thus, neuropeptide signaling through NK1R and NK2R may modulate the surface expression levels of HLA class II and costimulatory molecules on MoDCs, suggesting altered antigen presentation to CD4+ T cells. To confirm the effect of NK2R-mediated signaling on DC antigen presentation, we induced Bet-v1–specific IFN-γ– or IL-4–producing TH cells in vitro using white birch-derived antigen (see Fig E4 in this article's Online Repository at www.jacionline.org), as described previously.5Nagato T. Kobayashi H. Yanai M. Sato K. Aoki N. Oikawa K. et al.Functional analysis of birch pollen allergen Bet v 1-specific regulatory T cells.J Immunol. 2007; 178: 1189-1198Crossref PubMed Scopus (18) Google Scholar, 6Ohtake J. Ohkuri T. Togashi Y. Kitamura H. Okuno K. Nishimura T. Identification of novel helper epitope peptides of Survivin cancer-associated antigen applicable to developing helper/killer-hybrid epitope long peptide cancer vaccine.Immunol Lett. 2014; 161: 20-30Crossref PubMed Scopus (19) Google Scholar T-cell proliferation and cytokine production were evaluated after stimulation with antigen-loaded MoDCs. Blocking NK2R-mediated signaling suppressed the induction of antigen-specific CD4+ T cells (Fig 2, B). Furthermore, we confirmed the production of cytokines by ELISA for IL-2, IL-4, IL-5, IL-13, and IFN-γ by activated Bet-v1–specific TH cells after antigen stimulation by MoDCs. The production of TH1 and TH2 cytokines was decreased in the presence of NK1R and/or NK2R antagonists or by culturing with NK1R- or NK2R-siRNA–transfected MoDCs (Fig 2, C; see Fig E5, A, and Fig E6 in this article's Online Repository at www.jacionline.org). Furthermore, MoDC IL-12 production, as detected by ELISA in culture supernatants, was reduced in the presence of NK2R or NK1R antagonists (Fig 2, C). Previous reports demonstrated that DC-T-cell interactions through CD40-CD40L binding induces IL-12 production by DCs.7Kitamura H. Iwakabe K. Yahata T. Nishimura S. Ohta A. Ohmi Y. et al.The natural killer T (NKT) cell ligand alpha-galactosylceramide demonstrates its immunopotentiating effect by inducing interleukin (IL)-12 production by dendritic cells and IL-12 receptor expression on NKT cells.J Exp Med. 1999; 189: 1121-1128Crossref PubMed Scopus (563) Google Scholar It has also been shown that SP-NK1R signaling activates IL-12 secretion by DCs.8Janelsins B.M. Sumpter T.L. Tkacheva O.A. Rojas-Canales D.M. Erdos G. Mathers A.R. et al.Neurokinin-1 receptor agonists bias therapeutic dendritic cells to induce type 1 immunity by licensing host dendritic cells to produce IL-12.Blood. 2013; 121: 2923-2933Crossref PubMed Scopus (40) Google Scholar Thus, neuropeptide signaling through NK2R, as well as NK1R, promotes MoDC activation by antigen-specific TH cells. Finally, we confirmed that knockdown of NK1R and NK2R or the antagonists attenuated IFN-α, IFN-β, and IL-12p35 gene expression in MoDCs after poly I:C stimulation (see Fig E5, B, and Fig E7 in this article's Online Repository at www.jacionline.org). Notably, the combination of NK1R and NK2R antagonists synergistically inhibited cytokine production (see Fig E7). Taken together, these findings indicate that NK1R- or NK2R-dependent neuropeptide signaling activates antigen-specific TH cells by augmentation of DC function in both the induction and effector phases. This suggests that NK1R and NK2R antagonists may have both preventive and therapeutic effects in disorders induced by excessive DC/TH-mediated airway inflammation. Therefore, these findings may provide potential insights into the development of novel treatments for patients with severe inflammatory diseases caused by excessive type 1 immunity. Neuropeptides, such as SP and NKA, act mainly through their receptors, NK1R and NK2R, respectively, which may also be novel therapeutic targets in asthma. Previous studies demonstrated that antagonism of NK1R and NK2R by receptor antagonists reduced airway hyperresponsiveness and improved lung function.9Ramalho R. Soares R. Couto N. Moreira A. Tachykinin receptors antagonism for asthma: a systematic review.BMC Pulm Med. 2011; 11: 41Crossref PubMed Scopus (47) Google Scholar However, the clinical effects on airway inflammation and asthma symptoms are poorly understood. Recently, we found that human DCs upregulated the expression of TAC-1, which encodes SP and NKA, after IFN-γ or poly I:C stimulation.6Ohtake J. Ohkuri T. Togashi Y. Kitamura H. Okuno K. Nishimura T. Identification of novel helper epitope peptides of Survivin cancer-associated antigen applicable to developing helper/killer-hybrid epitope long peptide cancer vaccine.Immunol Lett. 2014; 161: 20-30Crossref PubMed Scopus (19) Google Scholar Thus, we speculated that type 1 immune-dominant severe asthma, which may be caused by viral infections, will be a promising target for therapy using NK1R and NK2R antagonists. We conclude that neuropeptide signaling through NK2R and NK1R promotes both innate and acquired immune responses through DC activation, suggesting that neuroimmune cross talk may be associated with various diseases, including infection and chronic inflammation such as severe asthma and HP. We thank Ms A. Nishiuchi for her excellent technical and secretarial assistance and Dr S. Ashino for thoughtful advice on this study. Fig E2View Large Image Figure ViewerDownload Hi-res image Download (PPT)Fig E3View Large Image Figure ViewerDownload Hi-res image Download (PPT)Fig E4View Large Image Figure ViewerDownload Hi-res image Download (PPT)Fig E5View Large Image Figure ViewerDownload Hi-res image Download (PPT)Fig E6View Large Image Figure ViewerDownload Hi-res image Download (PPT)Fig E7View Large Image Figure ViewerDownload Hi-res image Download (PPT) Download .doc (.07 MB) Help with doc files Online Repository Data Download .doc (.06 MB) Help with doc files Table E1 Download .doc (.03 MB) Help with doc files Table E2
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