Mechanisms of Cyclic Nucleotide Phosphodiesterases in Modulating T Cell Responses in Murine Graft-versus-Host Disease Michael Weber 1 , Corinna Lupp 1 , Pamela Stein 1 , Andreas Kreft 2 , Tobias Bopp 1 , Thomas C. Wehler 2 , Edgar Schmitt 1 , Hansjo ¨ rg Schild 1 , Markus P. Radsak 3 * 1 Institute for Immunology, Johannes Gutenberg-University Medical Center, Mainz, Germany, 2 Institute of Pathology, Johannes Gutenberg-University Medical Center, Mainz, Germany, 3 Department Internal of Medicine III, University Medical Center, Johannes Gutenberg-University, Mainz, Germany Abstract Graft-versus-host disease (GvHD) is a key contributor to the morbidity and mortality after allogeneic hematopoetic stem cell transplantation (HSCT). Regulatory Foxp3 + CD4 + T cells (T reg ) suppress conventional T cell activation and can control GvHD. In our previous work, we demonstrate that a basic mechanism of T reg mediated suppression occurs by the transfer of cyclic adenosine monophosphate (cAMP) to responder cells. Whether this mechanism is relevant for T reg mediated suppression of GvHD is currently unknown. To address this question, bone marrow and T cells from C57BL/6 mice were transferred into lethally irradiated BALB/c recipients, and the course of GvHD and survival were monitored. Transplanted recipients developed severe GvHD that was strongly ameliorated by the transfer of donor T reg cells. Towards the underlying mechanisms, in vitro studies revealed that T reg communicated with DCs via gap junctions, resulting in functional inactivation of DC by a metabolic pathway involving cAMP that is modulated by the phosphodiesterase (PDE) 4 inhibitor rolipram. PDE2 or PDE3 inhibitors as well as rolipram suppressed allogeneic T cell activation, indirectly by enhancing T reg mediated suppression of DC activation and directly by inhibiting responder T cell proliferation. In line with this, we observed a cooperative suppression of GvHD upon T reg transfer and additional rolipram treatment. In conclusion, we propose that an important pathway of T reg mediated control of GvHD is based on a cAMP dependent mechanism. These data provide the basis for future concepts to manipulate allogeneic T cell responses to prevent GvHD. Citation: Weber M, Lupp C, Stein P, Kreft A, Bopp T, et al. (2013) Mechanisms of Cyclic Nucleotide Phosphodiesterases in Modulating T Cell Responses in Murine Graft-versus-Host Disease. PLoS ONE 8(3): e58110. doi:10.1371/journal.pone.0058110 Editor: Christian Taube, Leiden University Medical Center, The Netherlands Received September 3, 2012; Accepted January 30, 2013; Published March 6, 2013 Copyright: ß 2013 Weber et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported by grants from the Deutsche Forschungsgemeinschaft to M.P.R. and H.S. (Ra988/4-2), SFB TR52 TPA1 (T.B. and E.S.), the GRK 1043 International Graduate School of Immunotherapy (E.S. and T.B.), ‘‘Forschungszentrum Immunologie (FZI)’’ of the University medical center (H.S., E.S. and T.B.). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: radsak@uni-mainz.de Introduction For patients with high risk hematological malignancies alloge- neic hematopoetic stem cell transplantation (HSCT) is the only curative treatment option [1]. The therapeutic principle of HSCT relies on a graft versus leukemia (GvL) or graft versus tumor (GvT) effect generated by donor lymphocytes that specifically recognize and eliminate malignant cells in the recipient [2]. However, after HSCT additional immune responses may occur against healthy tissues creating graft-versus-host disease (GvHD), an important contributor to transplant related morbidity and mortality [3]. To improve the feasibility of HSCT, it will be crucial to gain the ability to guide immune responses in the desired way maintaining anti-viral and anti-tumor responses while controlling undesired responses, namely GvHD. Naturally occuring regulatory T cells (T reg ) are responsible for maintaining peripheral self tolerance [4], but they may also play a role in the failure to control tumor growth as T reg cell depletion can facilitate tumor rejection [5,6]. In the context of HSCT, T reg cells have been shown to control GvHD [7–9], while on the other hand preserving GvT reactions [10]. However, the strategies to control effector T or T reg cell activity might be relevant since stringent inhibition of both may result in poor tumor outcome [9]. In addition, current clinical protocols demonstrate the feasibility and safety of T reg cell transfer in humans [11], possibly opening T reg based treatment options for patients beyond experimental settings in the near future. Hence, it is important to understand the relevant mechanisms of T reg mediated suppression in HSCT. Donor T cells are activated by conventional CD11c + dendritic cells (DC) in the HSCT recipient [12–14]. In this context, the blockade of costimulatory molecules induces transplantation tolerance that is mediated by T cell anergy or T reg subpopulations depending on the particular model used [15–17]. In general, T reg mediated suppression occurs by cytokine independent [18–20], but contact dependent ways, i. e. via the glucocorticoid induced tumor necrosis factor receptor (GITR) [20], CTLA-4 or mem- brane-bound TGF-b [21]. Beyond this, the transfer of cyclic adenosine monophosphate (cAMP) to target cells via gap junction intercellular communication (GJIC) is a key mechanism of T reg mediated suppression [22,23], also important in T reg -DC interac- tion that occurs by a direct contact (cAMP) dependent and by a contact independent pathway [24]. PLOS ONE | www.plosone.org 1 March 2013 | Volume 8 | Issue 3 | e58110