Phenotype and Effector Function of CC Chemokine Receptor 9-Expressing Lymphocytes in Small Intestinal Crohn's Disease This information is current as of September 29, 2021. Masayuki Saruta, Qi T. Yu, Armine Avanesyan, Phillip R. Fleshner, Stephan R. Targan and Konstantinos A. Papadakis J Immunol 2007; 178:3293-3300; ; doi: 10.4049/jimmunol.178.5.3293 http://www.jimmunol.org/content/178/5/3293 Downloaded from References This article cites 26 articles, 12 of which you can access for free at: http://www.jimmunol.org/content/178/5/3293.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 • Fast Publication! 4 weeks from acceptance to publication by guest on September 29, 2021 *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 © 2007 by The American Association of Immunologists All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology Phenotype and Effector Function of CC Chemokine Receptor 9-Expressing Lymphocytes in Small Intestinal Crohn’s Disease1 Masayuki Saruta,2*QiT.Yu,2* Armine Avanesyan,* Phillip R. Fleshner,† Stephan R. Targan,* and Konstantinos A. Papadakis3* CCL25/CCR9 chemokine ligand/receptor pair has been reported to play an important role in small bowel (SB) immunity and inflammation. We have previously reported an aberrant SB expression of CCL25 in Crohn’s disease (CD) and an increased frequency of CCR9؉ T cells in the peripheral blood of patients with SB inflammatory diseases such as CD and celiac disease. In .this study, we have characterized the phenotype and effector function of CCR9؉ T cells in mucosal lymphoid tissues in CD ؉ We show that CCR9 T cells isolated from mesenteric lymph nodes (MLN) draining CD SB express a more activated Downloaded from phenotype compared with MLN draining normal SB. Stimulation of CCR9؉ T cells isolated from CD SB lamina propria produced more IFN-␥ and IL-17 in response to anti-CD3 or IL-12/IL-18 stimulation compared with those isolated from normal SB. The addition of TL1A to the cytokine combination markedly augmented the secretion of IFN-␥, but not IL-17, by CD lamina propria CCR9؉ T cells. CCL25 incubation of CD SB lamina propria lymphocytes and MLN lymphocytes ␤increased their adhesion to VCAM-1/Fc in vitro. Finally, the TCRV␤ analysis of CCR9؉ T cells revealed a diverse TCRV ؉ ؉ repertoire among MLN CCR9 T cells in patients with SB CD. Our data indicate that CCR9 T cells in SB CD are http://www.jimmunol.org/ proinflammatory and support the rationale for the use of CCR9 antagonists for the treatment of human SB CD. The Journal of Immunology, 2007, 178: 3293–3300. rohn’s disease (CD)4 is a chronic, relapsing and remitting a critical role in immune homeostasis by regulating lymphocyte inflammatory intestinal disorder that can affect any part trafficking to or within lymphoid organs and in peripheral tissues C of the gastrointestinal tract but often involves the small (7–13). They are classified as C, CC, CXC, and CX3C based on bowel (SB), colon, or both (1). Although CARD15/NOD2 gene the positioning of cysteine residues that form two disulfide bonds variants are associated with ileal location of CD (2, 3), the path- (14). Chemokines mediate their actions through chemokine recep- by guest on September 29, 2021 ways of immune cell trafficking selectively to the SB in CD tors on the surface of target cells. Chemokine receptors are cur- have not been defined adequately. Recruitment of T lympho- rently divided into four families based on the type of chemokine cytes to the intestinal mucosa plays an important role in the that they bind; they are CXCR1 to CXCR6, CCR1 to CCR10, pathogenesis of inflammatory bowel disease (IBD). Chemo- XCR1, or CX3CR1 (15). kines and their receptors as well as adhesion molecules that are CCL25 is selectively expressed in the thymus and small intes- involved in regulating these processes represent novel thera- tine but not colon (9, 10). The only known receptor for CCL25, peutic targets for human IBD (4–6). CCR9, is highly expressed on developing thymocytes and small ϳ Chemokines constitute a large family of small ( 8–14 kDa), intestinal lamina propria lymphocytes (LPL) and intraepithelial structurally related heparin binding proteins, which are constitu- lymphocytes (9, 10, 16). We and others (9, 10, 12) have proposed tively expressed in lymphoid and extra lymphoid tissues, and play that the CCL25/CCR9 chemokine ligand receptor pair may play an important role in the regional specialization of intestinal immunity and that the coexpression of CCR9 and ␣ ␤ on the cell surface *Inflammatory Bowel Disease Center and Immunobiology Institute, and †Division of 4 7 Colorectal Surgery, Cedars-Sinai Medical Center and the UCLA School of Medicine, may provide a small intestinal “address code” for circulating in- Los Angeles, CA 90048 testinal memory T cells. We previously reported, that in peripheral Received for publication June 27, 2006. Accepted for publication December 27, 2006. blood (PB), CCR9ϩ T lymphocytes were markedly elevated in The costs of publication of this article were defrayed in part by the payment of page patients with SB immune-mediated diseases, including CD, but not charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. in patients with purely colonic CD, and that CCL25 expression is altered in inflamed SB but is not expressed in either normal or 1 This work was supported by grants from the Broad Medical Research Program in Inflammatory Bowel Diseases by the Eli and Edythe L. Broad Foundation (to K.A.P.). inflamed colon. These data suggest the potential involvement of ϩ 2 M.S. and Q.T.Y. contributed equally to this work and should be considered as first CCR9 T cells in the pathogenesis of SB CD (11). Furthermore, authors. we have demonstrated that the memory subset of circulating CCR9ϩ ϩ 3 Address correspondence and reprint requests to Dr. Konstantinos A. Papadakis, CD4 T cells in healthy donors has characteristics of mucosal T Cedars-Sinai Medical Center, 8700 Beverly Boulevard, D-4063, Los Angeles, CA 90048. E-mail address: [email protected] lymphocytes since they have an activated phenotype, respond to anti-CD2 stimulation, and exhibit a Th1 or T regulatory 1 cytokine 4 Abbreviations used in this paper: CD, Crohn’s disease; HPF, high power field; IBD, inflammatory bowel disease; LP, lamina propria; LPL, lamina propria lymphocyte; profiles (12). MFI, mean fluorescence intensity; MLN, mesenteric lymph node; PB, peripheral In this study, we have characterized the phenotype and effector blood; SB, small bowel; UC, ulcerative colitis. function of CCR9ϩ T cells in SB mucosal lymphoid tissues in Copyright © 2007 by The American Association of Immunologists, Inc. 0022-1767/07/$2.00 patients with SB CD and compared it with normal controls. We www.jimmunol.org 3294 CCR9ϩ T LYMPHOCYTES IN SMALL INTESTINAL CD Downloaded from http://www.jimmunol.org/ by guest on September 29, 2021 FIGURE 1. Phenotypic characteristics of CCR9ϩ T cells from normal and inflamed CD SB. A, LPL lymphocytes were isolated from normal and CD SB and stained for CCR9, CD3, and the indicated surface markers. The ϩ cells were gated on CD3 T cells and analyzed by FACS. The mean per- ϩ FIGURE 2. Phenotype of CCR9 T cells from MLN draining normal centage Ϯ SD of surface markers among CCR9ϩ T cells are shown be- and CD SB. A, MLN lymphocytes draining SB were isolated from five CD tween normal and CD SB (n ϭ 6). A representative FACS analysis of SB and three normal specimens and stained with CCR9, CD3, and the indi- LPL from CD (B) and normal SB (C) is shown. The numbers indicate the ϩ ϩ cated surface markers. The cells were gated on CD3 T cells and analyzed percentage of marker positive cells among CCR9 T cells (right upper ϩ by FACS. The mean percentage Ϯ SD of surface markers among CCR9 panel) and CCR9Ϫ T cells (right lower panel). T cells is shown. A representative FACS analysis of MLN draining CD SB (B) and normal SB (C) is shown. The numbers indicate the percentage of ϩ ϩ marker-positive cells among CCR9 T cells (right upper panel) and show that CCR9 T cells isolated from mesenteric lymph node CCR9Ϫ T cells (right lower panel). (MLN)-draining CD SB have an activated phenotype and those from CD SB lamina propria (LP) exhibit a predominant Th1 and ThIL-17 cytokine profile. CCR9ϩ T cells isolated from MLN- draining SB CD express a polyclonal TCRV␤ repertoire. In addi- Materials and Methods tion, we show that CCL25 triggers the adhesion of CD SB LPL and Patients MLN lymphocytes to VCAM-1/Fc in vitro and therefore could ϩ We enrolled patients with CD, ulcerative colitis (UC), or non-IBD gastro- contribute to the CCR9 T cell trafficking to the SB in inflamma- intestinal disorders who were hospitalized at the Cedars-Sinai Medical tory conditions. Center and underwent intestinal resection. The study was approved by the The Journal of Immunology 3295 (Sigma-Aldrich), 0.1 mg/ml DNase I (Sigma-Aldrich), 50 ␮g/ml gen- tamicin, 100 U/ml penicillin, 100 ␮g/ml streptomycin, and 50 ␮g/ml fungizone, in a shaking water bath (100 rpm).