Identification of Functional Roles for Both IL-17RB and IL-17RA in Mediating IL-25-Induced Activities This information is current as Erika A. Rickel, Lori A. Siegel, Bo-Rin Park Yoon, James B. of September 29, 2021. Rottman, David G. Kugler, David A. Swart, Penny M. Anders, Joel E. Tocker, Michael R. Comeau and Alison L. Budelsky J Immunol 2008; 181:4299-4310; ; doi: 10.4049/jimmunol.181.6.4299 Downloaded from http://www.jimmunol.org/content/181/6/4299 References This article cites 30 articles, 16 of which you can access for free at: http://www.jimmunol.org/content/181/6/4299.full#ref-list-1 http://www.jimmunol.org/ Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists by guest on September 29, 2021 • Fast Publication! 4 weeks from acceptance to publication *average Subscription Information about subscribing to The Journal of Immunology is online at: http://jimmunol.org/subscription Permissions Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html Email Alerts Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts The Journal of Immunology is published twice each month by The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 Copyright © 2008 by The American Association of Immunologists All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology Identification of Functional Roles for Both IL-17RB and IL-17RA in Mediating IL-25-Induced Activities Erika A. Rickel,* Lori A. Siegel,* Bo-Rin Park Yoon,* James B. Rottman,† David G. Kugler,* David A. Swart,* Penny M. Anders,* Joel E. Tocker,* Michael R. Comeau,* and Alison L. Budelsky1* IL-25 (IL-17E) is a unique IL-17 family ligand that promotes Th2-skewed inflammatory responses. Intranasal administration of IL-25 into naive mice induces pulmonary inflammation similar to that seen in patients with allergic asthma, including increases in bronchoalveolar lavage fluid eosinophils, bronchoalveolar lavage fluid IL-5 and IL-13 concentrations, goblet cell hyperplasia, and increased airway hyperresponsiveness. IL-25 has been reported to bind and signal through IL-17RB (IL-17BR, IL-17Rh1). It has been demonstrated recently that IL-17A signals through a heteromeric receptor composed of IL-17RA and IL-17RC. We sought to determine whether other IL-17 family ligands also utilize heteromeric receptor complexes. The required receptor Downloaded from subunits for IL-25 biological activities were investigated in vitro and in vivo using a combination of knockout (KO) mice and antagonistic Abs. Unlike wild-type mice, cultured splenocytes from either IL-17RB KO or IL-17RA KO mice did not produce IL-5 or IL-13 in response to IL-25 stimulation, and both IL-17RB KO and IL-17RA KO mice did not respond to intranasal admin- istration of IL-25. Furthermore, treatment with antagonistic mAbs to either IL-17RB or IL-17RA completely blocked IL-25- induced pulmonary inflammation and airway hyperresponsiveness in naive BALB/c mice, similar to the effects of an antagonistic http://www.jimmunol.org/ Ab to IL-25. Finally, a blocking Ab to human IL-17RA prevented IL-25 activity in a primary human cell-based assay. These data demonstrate for the first time that IL-25-mediated activities require both IL-17RB and IL-17RA and provide another example of an IL-17 family ligand that utilizes a heteromeric receptor complex. The Journal of Immunology, 2008, 181: 4299–4310. nterleukin-25 (also known as IL-17E) is a member of the a soluble IL-17RB.Fc protein or an antagonistic Ab to IL-25 in- structurally related IL-17 family of cytokines that were iden- hibits many of the components of the Ag-induced Th2-type pul- I tified based on sequence homology to IL-17A (1, 2). Unlike monary inflammation (9, 10). the proinflammatory effects associated with IL-17A, IL-25 pro- The biological effects of IL-25 are reported to be mediated motes Th2-type immune responses. For example, transgenic ex- through IL-17RB (also known as IL-17BR or IL-17Rh1), a mem- by guest on September 29, 2021 pression of IL-25 in mice results in increased blood eosinophilia, ber of the IL-17R family of cytokine receptors (1, 2). IL-25 binds serum IgE, IgG1, IL-13, and IL-5 concentrations and gene expres- with high affinity to and activates signaling pathways through IL- sion of IL-4, IL-5, IL-10, and IL-13 in multiple tissues (3, 4). In 17RB (11–14). It has, however, recently been reported that IL-17A contrast, IL-25 knockout (KO)2 mice exhibit an impaired Th2 re- and IL-17F signal through a heteromeric receptor complex con- sponse to infection by the parasitic helminthes Nippostrongylus taining IL-17RA and IL-17RC (15). In light of this information, brasiliensis and Trichuris muris, resulting in increased suscepti- we investigated the receptor subunits required for IL-25 activities bility to infection and chronic inflammation (5, 6). using a combination of KO mice and antagonistic Abs. The roles The effects of IL-25 specifically in the lung have also been in- for both IL-17RB and IL-17RA in mediating IL-25-induced events vestigated. Intranasal administration of either IL-25-expressing ad- were first investigated using IL-17RB KO and IL-17RA KO mice. enovirus or rIL-25 protein increases bronchoalveolar lavage fluid KO mouse experiments were validated by studies using antago- (BALF) and lung tissue eosinophilia, airway hyperresponsiveness, nistic Abs to these receptors. Finally, the mouse data were fol- mucus secretion, and lung mRNA levels of IL-4, IL-5, IL-13, and lowed up by examining the role for IL-17RA in IL-25-mediated CCL11/eotaxin (7, 8). In an OVA-induced pulmonary inflamma- activity in a human primary cell-based assay. Taken together, these tion model, IL-25 mRNA expression increases in the lung after data provide compelling evidence that IL-25 biological activities aerosolized OVA challenge, and antagonism of IL-25 using either require both IL-17RB and IL-17RA, identifying IL-25 as another IL-17 family cytokine that utilizes IL-17RA in a heteromeric re- *Department of Inflammation Research, Amgen, Seattle, WA 98119; and †Depart- ceptor complex. ment of Pathology, Amgen, Cambridge, MA 02139 Received for publication March 17, 2008. Accepted for publication July 20, 2008. The costs of publication of this article were defrayed in part by the payment of page Materials and Methods charges. This article must therefore be hereby marked advertisement in accordance Mice with 18 U.S.C. Section 1734 solely to indicate this fact. 1 Address correspondence and reprint requests to Dr. Alison Budelsky, Amgen, 1201 C57BL/6 and BALB/c wild-type (WT) mice were obtained from Charles Ϫ/Ϫ Amgen Court West, Seattle, WA 98119. E-mail address: [email protected] River Laboratories and Taconic Farms. The generation of IL-17RA 2 (IL-17RA KO) mice has been described previously (16), and the IL-17RA Abbreviations used in this paper: KO, knockout; WT, wild type; BALF, bronchoal- Ϫ/Ϫ veolar lavage fluid; BAL, bronchoalveolar lavage; AHR, airway hyperresponsiveness; KO mouse breeding colony is maintained at Taconic Farms. IL-17RB MCh, methacholine; PAS, periodic acid-Schiff; TSLP, thymic stromal lymphopoietin; (IL-17RB KO) mice were generated as follows. A gene targeting vector i.n. ϭ intranasal; PENH, enhanced pause; TRAF6, TNFR-associated factor 6. was constructed with a 2107-bp 5Ј arm of homology and a 5889-bp 3Ј arm of homology flanking a neomycin resistance cassette. A thymidine kinase Copyright © 2008 by The American Association of Immunologists, Inc. 0022-1767/08/$2.00 cassette was inserted into the 5Ј end of the vector. The targeting vector was www.jimmunol.org 4300 IL-17RB AND IL-17RA ARE REQUIRED FOR IL-25 ACTIVITIES designed to remove a region of exon 3 of the murine IL-17RB gene en- Intranasal cytokine challenge protocol coding aa H38-R73. 129-derived embryonic stem cells were electroporated Mice were intranasally dosed with 0.5 ␮g of murine IL-25 (Amgen), 0.5 with the targeting vector and selected in the presence of G418 and ganci- ␮ ␮ clovir as previously described (17). Embryonic stem cell clones carrying g of murine IL-13 (Biosystems), 0.5 g of murine IL-17F (R&D Sys- tems), or an equal volume (50 ␮l) of vehicle. Cytokines were diluted in the targeted insertion in IL-17RB were identified by a combination of PCR ␮ and genomic Southern blot analyses and were injected into Swiss Black PBS (Life Technologies) containing 10 g/ml mouse serum albumin blastocysts. PCR genotyping was performed using primers 3291-82 (5Ј- (Sigma-Aldrich) or in PBS alone. IL-25, IL-13, and IL-17F intranasal chal- CCAGGATCCAGGAGTTTAGCTT-3Ј) and 3291-83 (5Ј-CCCTATGTC lenge was performed once daily for 4 consecutive days. All mice were analyzed 24 h after the final intranasal challenge. CTTCCCGTTTCTA-3Ј) that produce a 336-bp product for the WT allele and a 280-bp product for the KO allele. Male chimeras were crossed to Ab treatment protocol Swiss Black females to generate mice heterozygous for the IL-17RB mu- tation (IL-17RBϩ/Ϫ). IL-17RBϩ/Ϫ mice were subsequently intercrossed to BALB/c mice were injected i.p. once daily for 4 consecutive days with 250 generate IL-17RBϪ/Ϫ mice (IL-17RB KO). IL-17RB KO mice were ␮g of anti-IL-25 mAb (Amgen), 250 ␮g of anti-IL-17RB mAb (Amgen), moved to a C57BL/6 background using Marker-Assisted Accelerated 200 ␮g of anti-IL-17RA-mAb (Amgen), 200 ␮g of anti-IL-17A mAb (Am- Backcrossing (MAX-BAXSM) technology at Charles River Laboratories.