CD69 Controls the Pathogenesis of Allergic Airway Inflammation 1 Takako Miki-Hosokawa, 2 * Akihiro Hasegawa, 2 * † Chiaki Iwamura, 2 * Kenta Shinoda,* Soichi Tofukuji,* Yukiko Watanabe,* Hiroyuki Hosokawa,* Shinichiro Motohashi,* Kahoko Hashimoto, ‡ Mutsunori Shirai, † Masakatsu Yamashita,* and Toshinori Nakayama 3 * Airway inflammation and airway hyperresponsiveness are central issues in the pathogenesis of asthma. CD69 is a membrane molecule transiently expressed on activated lymphocytes, and its selective expression in inflammatory infiltrates suggests that it plays a role in the pathogenesis of inflammatory diseases. In CD69-deficient mice, OVA-induced eosinophilic airway inflammation, mucus hyperproduction, and airway hyperresponsiveness were attenuated. Cell transfer of Ag-primed wild-type but not CD69- deficient CD4 T cells restored the induction of allergic inflammation in CD69-deficient mice, indicating a critical role of CD69 expressed on CD4 T cells. Th2 responses induced by CD69-deficient CD4 T cells in the lung were attenuated, and the migration of CD4 T cells into the asthmatic lung was severely compromised. The expression of VCAM-1 was also substantially altered, suggesting the involvement of VCAM-1 in the CD69-dependent migration of Th2 cells into the asthmatic lung. Interestingly, the administration of anti-CD69 Ab inhibited the induction of the OVA-induced airway inflammation and hyperresponsiveness. This inhibitory effect induced by the CD69 mAb was observed even after the airway challenge with OVA. These results indicate that CD69 plays a crucial role in the pathogenesis of allergen-induced eosinophilic airway inflammation and hyperresponsiveness and that CD69 could be a possible therapeutic target for asthmatic patients. The Journal of Immunology, 2009, 183: 8203– 8215. A sthma is a chronic inflammatory disease of the lower airways that causes airway hyperresponsiveness (AHR) 4 to a wide variety of specific and nonspecific stimuli (1, 2). In most cases, the extent of AHR correlates with the level of airway inflammation. Hallmarks of asthma include airway inflam- mation predominated by eosinophils, mucus hyperproduction, and Th2 cytokines (IL-4, IL-5, and IL-13) (3–7). A suggestion for a Th2 paradigm for allergic diseases, wherein increased activation of Th2 cells that produce Th2 cytokines results in IgE production and the recruitment and activation of eosinophils, comes from obser- vations of animal models previously studied. This notion has been supported by clinical studies in which the release of Th2-like cy- tokines from the lymphocytes of asthmatic patients was demon- strated (8, 9). CD69 is a type II membrane protein expressed as a homodimer composed of heavily glycosylated subunits (10). CD69 is known as an early activation marker Ag of lymphocytes (11, 12). Freshly prepared thymocytes undergoing selection events express CD69, and regulatory roles for CD69 expression in T cell development in the thymus have been suggested (13, 14). The regulatory roles of CD69 in a collagen-induced arthritis model (15) and an anti- collagen Ab-induced arthritis model (16) were reported and mul- tiple target processes were suggested; however, the role of CD69 in other inflammatory models, such as in the allergic airway in- flammation, has not been clarified. More recently, a new function of CD69 in the lymphocyte trafficking has been proposed (17). We herein investigated the role of CD69 using a mouse model of allergic asthma and found that CD69 plays a critical role in the induction of both Ag-induced eosinophilic airway inflammation and AHR. Furthermore, administration of anti-CD69 Ab resulted in a dramatic reduction in the extent of airway inflammation and AHR, suggesting that the CD69 mAb could be used for the treat- ment of asthmatic patients. Materials and Methods Mice CD69-deficient (CD69KO) mice (16) were backcrossed with BALB/c 10 times. BALB/c and C57BL/6 mice were purchased from Charles River Laboratories. GFP-transgenic (Tg) mice with a C57BL/6 background ex- pressed an enhanced GFP in all tissue under the control of the -actin promoter (18). All mice including OVA-specific -TCR-transgenic (DO11.10 Tg) mice (19) were maintained under specific pathogen-free conditions. All animal care was conducted in accordance with the guide- lines of Chiba University. *Department of Immunology, Graduate School of Medicine, Chiba University, Chiba, Japan; † Department of Microbiology and Immunology, Yamaguchi University School of Medicine, Yamaguchi, Japan; and ‡ Department of Life and Environmental Sci- ences and High Technology Research Center, Chiba Institute of Technology, Chiba Japan Received for publication February 26, 2009. Accepted for publication October 9, 2009. 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 the Global Center of Excellence Program (Global Center for Education and Research in Immune System Regulation and Treatment), MEXT (Monbukagakusho) (Japan), and by grants from the Ministry of Education, Culture, Sports, Science and Technology (Japan) (Grants-in-Aid for Scientific Re- search on Priority Areas nos. 17016010 and 20060003, Scientific Research (B) no. 17390139, Scientific Research (C) nos. 18590466, 19590491, 19591609, and 20590485, Exploratory Research no. 19659121, and Young Scientists (B) no. 20790367, (Start-up) 20890038: Special Coordination Funds for Promoting Science and Technology, and Cancer Translational Research Project), the Ministry of Health, Labor and Welfare (Japan), and The Japan Health Science Foundation. This work was also supported by the Naito Foundation. 2 T.M.-H., A.H., and C.I. contributed equally to this work. 3 Address correspondence and reprint request to Dr. Toshinori Nakayama, Depart- ment of Immunology, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8670 Japan. E-mail address: tnakayama@faculty.chiba-u.jp 4 Abbreviations used in this paper: AHR, airway hyperresponsiveness; BAL, bronchioalveolar lavage; Cdyn, dynamic compliance; PAS, periodic acid-Schiff; RL, lung resistance; S1P 1 , sphingosine 1 phosphate receptor 1; Tg, transgenic; KO, knockout; WT, wild type; HPRT, hypoxanthine phosphoribosyltransferase; Penh, enhanced pause. Copyright © 2009 by The American Association of Immunologists, Inc. 0022-1767/09/$2.00 The Journal of Immunology www.jimmunol.org/cgi/doi/10.4049/jimmunol.0900646