CCR8 Expression Identifies CD4 Memory T Cells Enriched for FOXP3  Regulatory and Th2 Effector Lymphocytes Dulce Soler, 1 Tobias R. Chapman, Louis R. Poisson, Lin Wang, Javier Cote-Sierra, Mark Ryan, Alice McDonald, Sunita Badola, Eric Fedyk, Anthony J. Coyle, 2 Martin R. Hodge, and Roland Kolbeck 3 CD4  Th2 cells are important regulators of allergic inflammation. CCR8 is thought to play a role in Th2-mediated responses, however, expression of CCR8 in peripheral blood has not been fully characterized. Using a fluorescent form of the ligand selective for CCR8 (F-CCL1), we identified the leukocytes expressing CCR8 in human, monkey, and mouse peripheral blood. CCR8 expression is primarily restricted to a subset of human CD4 memory T lymphocytes (15%). Approximately 40% of CCR8  CD4  T cells express Th2 cytokines IL-4 or IL-13 while 13% express the Th1 cytokine IFN-. In fact, 50% of all Th2, but only 5% of Th1, cells express CCR8. Upon anti-CD3/anti-CD28 mAb-mediated activation, CCR8  CD4  T cells secrete 3- to 7-fold higher levels of IL-4, IL-5, IL-9, and IL-13 and 10- to 20-fold lower levels of IFN- or IL-17, compared with CCR8  CD4  memory T cells. Two-thirds of CCR8  CD4 T cells express cutaneous lymphocyte-associated Ag while the majority lack gut-homing receptors. CCR8  CD4  cells express CCR7 and CD62L and are present in spleen and lymph nodes of mice. Approximately 25% of CCR8  CD4 T cells express CD25 high while 20% of CCR8  CD4  express the T regulatory cell transcription factor FOXP3 accounting for 60% of all FOXP3-expressing CD4  T cells. In conclusion, CCR8 marks a diverse subset of CD4 memory T cells enriched for T regulatory and Th2 cells which have the potential for recruitment into sites of allergic inflammation where they could participate in the induction and regulation of the allergic response. The Journal of Immunology, 2006, 177: 6940 – 6951. T he T lymphocyte pool consists of naive T cells and Ag- experienced memory T cells which can be further divided into nonpolarized (T NPM ), 4 also referred to as central memory (T CM ), and effector memory (T EM ) T cells. T NPM expand and acquire T-effector functions in response to Ag re-encounter while T EM represent a circulating pool of polarized T cells capable of producing immediate effector functions upon restimulation. CD4 + T EM cells include Th1 and Th2 cells which migrate to in- flamed peripheral tissues where they secrete effector cytokines (IFN- by Th1 and IL-4, IL-5, IL-9, and IL-13 by Th2 cells, re- spectively) involved in amplifying the immune response (1, 2). In recent years, other populations of memory T cells that have the capacity to down-regulate immune responses through either cell- to-cell contact or the release of soluble mediators such as IL-10 and TGF-, so-called regulatory T (T REG ) cells, have gained in- creased attention (3). A subpopulation of T REG cells, known as naturally occurring T REG , are generated in the thymus during T cell development and are best characterized by high expression levels of the IL-2R -chain (CD25) and by the expression of the forkhead family transcription factor FOXP3 which is necessary for their development and function (4, 5). Migration of diverse T cell subsets during homeostasis and in- flammation is controlled by the concerted expression of adhesion molecules, such as integrins and selectins, and chemokine recep- tors. For example, coexpression of L-selectin (CD62L) and the chemokine receptor CCR7 by T cells is a prerequisite for homing to lymphoid tissues (6 – 8) while expression of the integrin  4  7 or cutaneous lymphocyte-associated Ag (CLA) are required for lymphocyte migration to the gastrointestinal tract or the skin, respectively (9 –11). Chemokine receptors belong to the class of seven transmem- brane G protein-coupled receptors and have been shown to medi- ate a variety of biological processes upon chemokine binding, including angiogenesis, leukocyte activation, and chemokine- induced transendothelial migration through integrin activation and subsequent transmigration (12). Chemokines are small secreted proteins (8 kDa) which can be divided into four subfamilies based on the spacing of two conserved cysteine residues (12). They can also be distinguished by their pattern of regulation: 1) lym- phoid chemokines such as CCL19, CCL21, and CXCL13 are con- stitutively expressed in lymphoid tissues and mediate the migra- tion of leukocytes into and within lymphoid tissues by engagement of their respective receptors CCR7 and CXCR5 (6, 13, 14), 2) chemokines constitutively expressed in nonlymphoid tissues, for example, CCL1 in the skin which attracts CCR8 + T cells (15), and 3) inducible chemokines such as CCL11 (16), CCL17 (17, 18), and CXCL10 (19) attract effector cells into inflamed tissues through engagement of CCR3, CCR4, and CXCR3, respectively. We and others have recently reported the expression of the che- mokine CCL1 by IgE-activated mast cells in vitro and in vivo, implicating CCL1 in the recruitment of inflammatory cell types involved in allergic inflammation (Refs. 20 –22 and J. A. Gonzalo, Y. Qiu, J. M. Lora, A. Al-Garawi, J. L. Villeval, J. Boyce, C. Inflammation, Millennium Pharmaceuticals, Cambridge, MA 02139 Received for publication February 10, 2006. Accepted for publication August 28, 2006. 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 Address correspondence and reprint requests to Dr. Dulce Soler, Millennium Phar- maceuticals, 35 Landsdowne Street, Cambridge, MA 02139. E-mail address: dsoler@mpi.com 2 Current address: MedImmune, One MedImmune Way, Gaithersburg, MD 20878. 3 Current address: Peptimmune, 64 Sidney Street, Cambridge, MA 02139. 4 Abbreviations used in this paper: T NPM , nonpolarized memory T cell; T CM , central memory T cell; T EM , effector memory T cell; T REG , regulatory T cell; CLA, cuta- neous lymphoid-associated Ag; F-CCL1, fluorescently labeled CCL-1; DAPI, 4',6'-diamidino-2-phenylindole. The Journal of Immunology Copyright © 2006 by The American Association of Immunologists, Inc. 0022-1767/06/$02.00