Cell extrinsic regulation by CTLA-4 has been strongly linked to Treg-cell populations, with increased levels of CTLA-4 Saracatinib solubility dmso message in Treg cells relative to other CD4+ T-cell types, and CTLA-4 expression required for effective Treg-cell function [11–15, 19]. Analyses of CTLA-4 levels in Treg cells have previously been limited to methods that do not discriminate between the isoforms, with the assumption that all the CTLA-4 detected, and thus all of the regulatory function mediated by it, arises solely from the receptor isoform of the molecule. Here, we demonstrate that human Treg-cell populations can also express sCTLA-4 prominently and

that, under some circumstances, it can contribute to their suppressive function. As might be expected, sCTLA-4 was shown to be redundant when conditions in vitro favor Teff-cell inhibition mediated by cell contact-dependent Treg-cell mechanisms [47]. Instead, the data indicate a model whereby sCTLA-4 is important for suppression when Treg-cell numbers are too few for effective direct cell contacts to be made. It should be noted that although Treg cells appear to be important in producing sCTLA-4, our study does not rule out additional sources such as other T-cell types, and sCTLA-4 transcripts have also been detected by qPCR in both monocytes and immature DCs [48]. Recently, a role of

selleck kinase inhibitor sCTLA-4 in murine Treg-cell function has been supported by targeted and selective knockdown of the soluble isoform, using the posttranscriptional silencing mechanism of RNA interference (RNAi) [49]. In that study, knockdown of the sCTLA-4 isoform in NOD mice gave rise to Treg cells that failed to inhibit colitis induced by transfer of CD4+CD45RBhi cells and also accelerated onset of diabetes. Our results using isoform-specific Ab blockade are complementary, and show that sCTLA-4 has inhibitory effects on murine

T-cell responses in vitro and may promote tumor spread in vivo in a model of metastatic melanoma. Indeed, in this model, the protective effects of selective sCTLA-4 and pan-specific anti-CTLA-4 antibodies were similar, suggesting a dominant role for the soluble isoform. Taken together, we provide a new model to explain the seemingly paradoxical nature of CTLA-4 activity in terms of its ability, PD-1 inhibiton both to provide intrinsic T-cell negative costimulation, and to regulate effector T-cell responses extrinsically. Instead of these dual functions being mediated solely by mCTLA-4, we propose a major contribution to extrinsic regulation by sCTLA-4. Blood samples were collected by venepuncture from healthy volunteer donors. The Grampian Health Board and the University of Aberdeen Ethical Committee approved investigation protocols. PBMCs were prepared and cultured essentially as previously described [50] in RPMI 1640 medium (Invitrogen, Paisley, UK) supplemented with 5% autologous human serum in an atmosphere of 37°C, 5% CO2.