FOXP3  CD4  CD25  Adaptive Regulatory T Cells Express Cyclooxygenase-2 and Suppress Effector T Cells by a Prostaglandin E 2 -Dependent Mechanism 1 Milada Mahic,* Sheraz Yaqub,* C. Christian Johansson, 2 * Kjetil Taske ´n, 3 * and Einar M. Aandahl* † CD4  CD25  regulatory T (T R ) cells suppress effector T cells by partly unknown mechanisms. In this study, we describe a population of human suppressive CD4  CD25  adaptive T R (T R adapt ) cells induced in vitro that express cyclooxygenase 2 (COX-2) and the transcription factor FOXP3. T R adapt cells produce PGE 2 and suppress effector T cell responses in a manner that is reversed by COX inhibitors and PGE 2 receptor-specific antagonists. In resting CD4  CD25  T cells, treatment with PGE 2 induced FOXP3 expression. Thus, autocrine and paracrine effects of PGE 2 produced by COX-2-positive T R adapt cells may be responsible for both the FOXP3  phenotype and the mechanism used by these cells to suppress effector T cells. The Journal of Immunology, 2006, 177: 246 –254. T he CD4 + CD25 + regulatory T (T R ) 4 cells are a unique population of T cells that maintain peripheral immune tolerance and inhibit autoreactive T cells (1–3). Although T cells with regulatory or suppressive properties are not strictly confined to the CD4 + CD25 + T cell compartment, T R cells are now well characterized in both mice and humans and play an im- portant role in various clinical conditions. T R cells inhibit allo- reactive T cells and suppress transplant rejection reactions, and the role of T R cells in cancer and chronic infectious diseases is a sub- ject of intense investigation (4). Through in vitro experiments, T R cells are shown to effectively inhibit effector T cell responses such as cytokine production and proliferation (5). At least two subpopulations of T R cells exist named naturally occurring T R (T R nat ) cells and adaptive T R (T R adapt ) cells (4, 6). The lineage relationship between these two subsets remains un- clear. In mice, T R nat cells are generated in the thymus by recog- nizing self-peptides with high avidity yet escaping negative selec- tion (7). T R nat cells play a crucial function in the normal immune system by suppressing autoreactive T cells and maintaining im- mune tolerance; however, suppressive T R cells also inhibit im- mune responses directed against infectious agents and neoplasms. Although T R cells suppress responding T cells in an Ag-nonspe- cific manner, T R cells need to be activated through the TCR to achieve suppressive activity (8). Thus, it is likely that T R cells specific for foreign Ags, termed T R adapt cells, are of extrathymic origin and generated in the periphery from the peripheral T cell repertoire. The possibility that T R cells can be generated in the periphery is supported by several in vitro and in vivo models dem- onstrating the induction of T R adapt cells from CD25 - T cells by prolonged or repeated antigenic stimulation (9, 10). Alternatively, differentiation of T R adapt cells can be induced by IL-10 or TGF- (11, 12). However, it is still not determined whether T R nat and T R adapt cells represent distinct lineages or just phenotypic variants induced at different anatomical sites. The transcription factor FOXP3 that is essential for the suppressive activity of T R nat cells was, in mouse models, initially thought to be a specific marker of T R nat cells that could not be induced in activated peripheral T cells (13–15). However, later studies have demonstrated induction of FOXP3 expression in T R adapt cells in both in vitro and in vivo models (16 –20). The mechanism of suppression of T effector cells by T R cells is not fully known. Transwell experiments support a contact-depen- dent mechanism for T R nat cells, although a short-range humoral factor with limited water solubility cannot be ruled out. A role for IL-10, TGF-, and CTLA-4 have been demonstrated for both T R nat cells and T R adapt cells, but it is not finally determined whether these factors primarily play a role in the differentiation process or in the suppressive activity (3). PGs have strong immunomodulatory activity within the immune system and can have both systemic and short-range autocrine and paracrine effects (21). PGE 2 binds to the G protein-coupled recep- tors EP2 and EP4 and effectively suppress T cell immune responses by eliciting a cAMP (protein kinase A) Csk inhibitory pathway lo- calized to lipid rafts (22–29). We have recently described the role of PGE 2 in a mouse model for retroviral-induced immunodeficiency syndrome (MAIDS) (30), and PGE 2 also suppresses autoimmune manifestations and allograft rejections in mouse and human studies (23, 31–33). Within the immune system, cyclooxygenase type 2 (COX-2) is induced during inflammatory reactions and is responsible for production of PGs and thromboxanes from arachidonic acid. *The Biotechnology Centre, and † Department of Gastroenterological Surgery, Ull- evaal University Hospital, University of Oslo, Oslo, Norway Received for publication December 6, 2005. Accepted for publication April 11, 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 This work was supported by grants from the National Programme for Research in Functional Genomics in Norway and a Cancer Programme grant in the Research Council of Norway, the Norwegian Cancer Society, Novo Nordic Foundation Com- mittee, and the European Union (Research and Technological Development Grant QLK3-CT-2002-02149). 2 Present address: Immune and Gene Therapy Unit, Cancer Center Karolinska, Karo- linska Institute, SE-171 77 Stockholm, Sweden. 3 Address correspondence and reprint requests to Dr. Kjetil Taske ´n, The Biotechnol- ogy Centre, University of Oslo, P.O. Box 1125, N-0317 Oslo, Norway. E-mail ad- dress: kjetil.tasken@biotek.uio.no 4 Abbreviations used in this paper: T R , regulatory T; T R nat , naturally occurring T R ; T R adapt , adaptive T R ; MAIDS, murine acquired immunodeficiency syndrome; COX-2, cyclooxygenase type 2; SEB, staphylococcal enterotoxin B; PKC, protein kinase C; IBMX, 3-isobutyl-1-methylxanthine. 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