CC Chemokine Receptor 9 Expression Defines a Subset of Peripheral Blood Lymphocytes with Mucosal T Cell Phenotype and Th1 or T-Regulatory 1 Cytokine Profile 1 Konstantinos A. Papadakis, 2 * Carol Landers,* John Prehn,* Elias A. Kouroumalis, † Sofia T. Moreno,* Jose-Carlos Gutierrez-Ramos, ‡ Martin R. Hodge, ‡ and Stephan R. Targan 2 * The chemokine receptor CCR9 is expressed on most small intestinal lamina propria and intraepithelial lymphocytes and on a small subset of peripheral blood lymphocytes. CCR9-expressing lymphocytes may play an important role in small bowel immunity and inflammation. We studied the phenotype and functional characteristics of CCR9  lymphocytes in blood from normal donors. A subset of CCR9  T cells have a phenotype of activated cells and constitutively express the costimulatory molecules CD40L and OX-40. In contrast to CCR9  , CCR9  CD4  peripheral blood T cells proliferate to anti-CD3 or anti-CD2 stimulation and produce high levels of IFN- and IL-10. IL-10-producing cells were exclusively detected within the CCR9  subset of CD4  T cells by intracellular staining and were distinct from IL-2- and IFN--producing cells. Moreover, memory CCR9  CD4  lymphocytes respond to CD2 stimulation with proliferation and IFN-/IL-10 production, whereas memory CCR9  CD4  cells were unrespon- sive. In addition, memory CCR9  CD4  T cells support Ig production by cocultured CD19  B cells in the absence of prior T cell activation or addition of exogenous cytokines. Our data show that the memory subset of circulating CCR9  CD4  T cells has characteristics of mucosal T lymphocytes and contains cells with either Th1 or T-regulatory 1 cytokine profiles. Studies on the cytokine profile and Ag specificity of this cell subset could provide important insight into small intestinal immune-mediated diseases and oral tolerance in humans. The Journal of Immunology, 2003, 171: 159 –165. C hemokines and their receptors play a critical role in the migration of different types of leukocytes, including na- ive and memory/effector T lymphocytes, into microana- tomic compartments of lymphoid organs and peripheral tissues (1–5). T cells in peripheral blood (PB) 3 express a diverse set of chemokine receptors, and studies in the last few years have sup- ported the notion that the patterns of chemokine receptors charac- terize functional T cell subpopulations. Naive T cells, which pref- erentially migrate to lymph nodes and Peyer’s patches, express CXCR4, the receptor for stromal cell-derived factor1/CXC ligand 12, and CCR7, the receptor for Epstein-Barr virus-induced mole- cule1 ligand chemokine/C-C chemokine ligand (CCL) 19, and sec- ondary lymphoid organ chemokine/CCL21. Mice deficient in CCR7 or secondary lymphoid organ chemokine/CCL21 have de- fective homing of naive T cells to secondary lymphoid organs (6, 7). Based on the expression of CCR7, memory T cells can be further distinguished into CCR7  central memory T cells with the potential to home to lymphoid organs and CCR7-effector memory T cells with the potential to migrate to inflamed tissue (8). CXCR5 expression on memory T cells defines a subset that localizes to B cell follicles and germinal centers and supports Ig production by cocultured B cells (9, 10), whereas CCR4 and CCR8 are highly expressed by human CD4  CD25  -regulatory T cells (11). Other chemokine receptors are preferentially expressed by different Th subsets; CCR3 is expressed predominantly by Th2 cells, and CCR5 or CXCR3 are expressed by Th1 cells (12–15). Expression of other chemokine receptors by peripheral blood (PB) T cells may define a tissue-specific homing potential, such as CCR4 or CCR10, which are preferentially expressed by cutaneous lym- phocyte Ag  -skin homing T cells (16, 17), and CCR9, which is expressed by  4  7  -intestine-homing lymphocytes (18 –21). These data suggest an important role of certain chemokines and their receptors in regulating the homing of specific T cell sub- sets into microanatomic compartments of lymphoid organs and peripheral tissues (4). We and others have proposed that the chemokine thymus-ex- pressed chemokine (TECK)/CCL25 and its receptor CCR9 may play an important role in the regional specialization of intestinal immunity and that the combined expression of CCR9 and  4  7 on the cell surface may provide a small intestinal address code for circulating intestinal memory T cells (20, 21). CCR9 is expressed on a small subset of PB CD4  (2– 4%) and CD8  T cells, most of which coexpress the mucosal homing ligand  4  7 (18). Here, we show that circulating CCR9  CD4  T cells from normal donors have characteristics of mucosal T cells in terms of an activated phenotype, proliferative response to anti-CD2 stimulation, a Th1 or T-regulatory 1 (Tr1) cytokine profile, and support for Ig pro- duction by cocultured B cells. This T cell subset may provide a peripheral window to small intestinal immunity in humans and a tool to study Ag specificity and cytokine profile of effector and *Burns and Allen Research Institute and Inflammatory Bowel Disease Center, Ce- dars-Sinai Medical Center, David Geffen School of Medicine at UCLA, Los Angeles, CA 90048; † Univerity Hospital of Heraklion, University of Crete, Heraklion, Crete, Greece; and ‡ Millennium Pharmaceuticals, Cambridge, MA 02139 Received for publication December 20, 2002. Accepted for publication April 18, 2003. 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 is supported by Grants DK-46763 and DK-56328 from the National Institutes of Health (to S.R.T.) and by a Career Development Award from the Crohn’s and Colitis Foundation of America (to K.A.P.). 2 Address correspondence and reprint requests to Dr. Konstantinos A. Papadakis, Cedars-Sinai Medical Center, 8700 Beverly Boulevard, D-4063, Los Angeles, CA 90048. E-mail address: Papadakisk@cshs.org or Dr. Stephan R. Targan, 8700 Beverly Boulevard, D-4063, Los Angeles, CA 90048. E-mail address: Targans@cshs.org 3 Abbreviations used in this paper: PB, peripheral blood; CCL, C-C chemokine li- gand; TECK, thymus-expressed chemokine; Tr1, T-regulatory 1; TC, tricolor. The Journal of Immunology Copyright © 2003 by The American Association of Immunologists, Inc. 0022-1767/03/$02.00