Activation-Induced FoxP3 Expression Regulates Cytokine Production in Conventional T Cells Stimulated with Autologous Dendritic Cells

Abstract

A defining feature of dendritic cells (DCs) is their ability to induce the proliferation of autologous T cells in the absence of foreign antigen-a process termed the "autologous mixed leukocyte reaction" (AMLR). We report that equine monocytederived DCs, but not macrophages, are potent inducers of the AMLR. The response is contact dependent and major histocompatibility complex class II dependent and primarily involves CD3؉ CD4 ؉ CD8 ؊ T cells. Upon stimulation with DCs or the mitogen concanavalin A, a subset of the proliferating T cells expresses the regulatory T-cell (Treg) transcription factor FoxP3. Although many of these FoxP3 ؉ T cells are capable of producing the effector cytokines interleukin-4 (IL-4) and gamma interferon (IFN-␥), they are more likely to produce IL-10 and less likely to produce IFN-␥ than equivalent FoxP3 DCs take up antigen from the peripheral tissues, process it, and present it to naive T cells in the draining lymph node. Depending on the local cytokine environment during DC-mediated T cell activation, the antigen-specific T cell can proliferate and differentiate into one of a variety of effector T cell phenotypes, including (i) gamma interferon (IFN-␥)-producing Th1 cells that are important for immunity to intracellular pathogens and tumors; (ii) interleukin-4 (IL-4)-, IL-5-, and IL-13-producing Th2 cells that promote antibody production, help prevent parasitic diseases, and are involved in allergic responses; and (iii) IL-17-producing Th17 cells that protect against extracellular infections (58). DCs also play a critical role in immune regulation because they are capable of both expanding thymus-derived FoxP3 ϩ (designated FOXP3 in humans) natural regulatory T cells (nTregs) and inducing naive T cells to develop into induced regulatory T cells (iTregs) in the periphery (55, 56). In turn, Tregs use a variety of mechanisms (such as the production of the anti-inflammatory cytokines IL-10 and transforming growth factor ␤ [TGF-␤]) to keep effector responses in check and to prevent immune-mediated disease (7).DCs also possess the unique ability to efficiently induce proliferation of autologous T cells in the absence of exogenous antigen. This process was first demonstrated with murine DCs several decades ago and was termed the "autologous mixed leukocyte reaction" (AMLR) (39). The AMLR has since been described with human cells and is thought to represent polyclonal activation of autoreactive T cells specific for self-antigens presented by DCs (2,3,10,38,44). Interestingly, the AMLR displays characteristics of a normal immune response (including specificity and memory) and is reduced in a variety of human disease states (23, 53). Furthermore, the self-reactive T cells in the AMLR demonstrate a capacity for immunosuppression and increased transcription of the Treg transcription factor FOXP3, suggesting that such DC-stimulated T cells are involved in immune regulation during the normal immune response in vivo (25,45,51).A protocol for generating monocyte-derived DCs has been described in the h...

Apoptotic cell capture by DCs induces unexpectedly robust autologous CD4+ T‐cell responses

“…We wondered whether these CD4 + T cells had suppressive properties. Indeed, previous studies have shown that self‐reactive T cells in the autologous mixed leukocyte reaction may be immunosuppressive .…”

“…DCs also possess the unique ability to induce proliferation of autologous T cells in the absence of exogenous antigen, a process termed “autologous mixed leukocyte reaction” . T cells activated in this process display normal immune response in terms of specificity and memory, and a capacity for immunosuppression . There is evidence that, in this process, DCs present peptides derived from self‐proteins , mostly derived from residual apoptotic cells .…”

Donor‐derived equine mesenchymal stem cells suppress proliferation of mismatched lymphocytes

“…There are several ways to stimulate T lymphocytes, namely binding antibodies to the T cell receptor‐specific antigen CD3 and CD28, mixed leucocyte reaction or using mitogens, such as PHA or concanavalin A . Due to the lack of antibodies against equine epitopes , T cell stimulation in horses has usually been carried out using mitogens . Proliferation of T cells using mitogens has already been demonstrated in man, mice and dogs .…”

Inflammatory licensed equine MSCs are chondroprotective and exhibit enhanced immunomodulation in an inflammatory environment

“…To generate inflammatory macrophages, monocytes were isolated according to previously described protocols [ 22 , 24 ]. Using MACS selection, CD14-positive (1:100 dilution, mouse anti-equine CD14 antibody, clone 105; courtesy of Dr Bettina Wagner, Cornell University, Ithaca, NY, USA) cells were isolated, counted, resuspended in macrophage media (DMEM containing 10% normal horse serum (NHS; HyClone Laboratories, Logan, UT, USA), l -glutamine (2 mM), penicillin (100 units/ml), and streptomycin (100 μg/ml)), and plated at 1.8 × 10 6 cells per well in six-well plates.…”

“…Using MACS selection, CD14-positive (1:100 dilution, mouse anti-equine CD14 antibody, clone 105; courtesy of Dr Bettina Wagner, Cornell University, Ithaca, NY, USA) cells were isolated, counted, resuspended in macrophage media (DMEM containing 10% normal horse serum (NHS; HyClone Laboratories, Logan, UT, USA), l -glutamine (2 mM), penicillin (100 units/ml), and streptomycin (100 μg/ml)), and plated at 1.8 × 10 6 cells per well in six-well plates. Horse serum was only used in macrophage culture to provide species-specific serum commonly used in macrophage culture experiments [ 24 ]. Macrophage media were added every 48 h during culture.…”