Key role of macrophages in tolerance induction via T regulatory type 1 (Tr1) cells.

T regulatory type 1 (Tr1) cells are a class of regulatory T cells (Tregs ) participating in peripheral tolerance, hence the rationale behind their testing in clinical trials in different disease settings. One of their applications is tolerance induction to allogeneic islets for long-term diabetes-free survival. Currently the cellular and molecular mechanisms that promote Tr1-cell induction in vivo remain poorly understood. We employed a mouse model of transplant tolerance where treatment with granulocyte colony-stimulating factor (G-CSF)/rapamycin induces permanent engraftment of allogeneic pancreatic islets in C57BL/6 mice via Tr1 cells. The innate composition of graft and spleen cells in tolerant mice was analyzed by flow cytometry. Graft phagocytic cells were co-cultured with CD4+ T cells in vitro to test their ability to induce Tr1-cell induction. Graft phagocytic cells were depleted in vivo at different time-points during G-CSF/rapamycin treatment, to identify their role in Tr1-cell induction and consequently in graft survival. In the spleen, the site of Tr1-cell induction, no differences in the frequencies of macrophages or dendritic cells (DC) were observed. In the graft, the site of antigen uptake, a high proportion of macrophages and not DC was detected in tolerant but not in rejecting mice. Graft-infiltrating macrophages of G-CSF/rapamycin-treated mice had an M2 phenotype, characterized by higher CD206 expression and interleukin (IL)-10 production, whereas splenic macrophages only had an increased CD206 expression. Graft-infiltrating cells from G-CSF/rapamycin-treated mice-induced Tr1-cell expansion in vitro. Furthermore, Tr1-cell induction was perturbed upon in-vivo depletion of phagocytic cells, early and not late during treatment, leading to graft loss suggesting that macrophages play a key role in tolerance induction mediated by Tr1 cells. Taken together, in this mouse model of Tr1-cell induced tolerance to allogeneic islets, M2 macrophages infiltrating the graft upon G-CSF/rapamycin treatment are key for Tr1-cell induction. This work provides mechanistic insight into pharmacologically induced Tr1-cell expansion in vivo in this stringent model of allogeneic transplantation.

Evidence for a pathogenic role of extrafollicular, IL-10-producing CCR6+B helper T cells in systemic lupus erythematosus.

Interleukin 10 (IL-10) is an antiinflammatory cytokine, but also promotes B cell responses and plays a pathogenic role in systemic lupus erythematosus (SLE). CD4+CCR6+IL-7R+T cells from human tonsils produced IL-10 following stimulation by naïve B cells, which promoted B cell immunoglobulin G (IgG) production. These tonsillar CCR6+B helper T cells were phenotypically distinct from follicular helper T (TFH) cells and lacked BCL6 expression. In peripheral blood, a CCR6+T cell population with similar characteristics was identified, which lacked Th17- and TFH-associated gene signatures and differentiation-associated surface markers. CD4+CCR6+T cells expressing IL-10, but not IL-17, were also detectable in the spleens of cytokine reporter mice. They provided help for IgG production in vivo, and expanded systemically in pristane-induced lupus-like disease. In SLE patients, CD4+CCR6+IL-7R+T cells were associated with the presence of pathogenic anti-dsDNA (double-stranded DNA) antibodies, and provided spontaneous help for autoantibody production ex vivo. Strikingly, IL-10-producing CCR6+T cells were highly abundant in lymph nodes of SLE patients, and colocalized with B cells at the margins of follicles. In conclusion, we identified a previously uncharacterized population of extrafollicular B helper T cells, which produced IL-10 and could play a prominent pathogenic role in SLE.

HRG/HER2/HER3 signaling promotes AhR-mediated Memo-1 expression and migration in colorectal cancer.

Colorectal cancer (CRC) is a complex disease with still unsatisfactory prognosis even in western societies, although substantial progress has been made in pre-screening programs, surgical techniques and targeted therapy options. Mediator of motility-1 (Memo-1) was previously recognized as an important effector of cell migration downstream of receptor tyrosine kinase signaling in breast cancer. This study identified Memo-1 as frequently overexpressed in CRC and established a close link between extracellular HER2 activation, AhR/ARNT transcriptional activity and Memo-1 expression. Dissection of the hMemo-1 gene promoter using reporter assays and chromatin IP techniques revealed recruitment of Aryl hydrocarbon receptor (AhR)/Aryl hydrocarbon receptor nuclear-translocator (ARNT) complex, which positively influenced Memo-1 expression in cancer cells. We found that Memo-1 depletion negatively influenced the cellular actin network and that its expression is required for HER2-mediated cell migration and invasion. Moreover, analyses of Memo-1 expression in primary CRC revealed correlation with clinical parameters that point to Memo-1 as a new prognostic factor of aggressive disease in CRC patients. Altogether, these observations demonstrate that Memo-1 is an important downstream regulator of HER2-driven CRC cell migration and invasion through connecting extracellular signals from membrane to the cytoskeletal actin network.

Transplant tolerance to pancreatic islets is initiated in the graft and sustained in the spleen.

The immune system is comprised of several CD4(+) T regulatory (Treg) cell types, of which two, the Foxp3(+) Treg and T regulatory type 1 (Tr1) cells, have frequently been associated with transplant tolerance. However, whether and how these two Treg-cell types synergize to promote allograft tolerance remains unknown. We previously developed a mouse model of allogeneic transplantation in which a specific immunomodulatory treatment leads to transplant tolerance through both Foxp3(+) Treg and Tr1 cells. Here, we show that Foxp3(+) Treg cells exert their regulatory function within the allograft and initiate engraftment locally and in a non-antigen (Ag) specific manner. Whereas CD4(+) CD25(-) T cells, which contain Tr1 cells, act from the spleen and are key to the maintenance of long-term tolerance. Importantly, the role of Foxp3(+) Treg and Tr1 cells is not redundant once they are simultaneously expanded/induced in the same host. Moreover, our data show that long-term tolerance induced by Foxp3(+) Treg-cell transfer is sustained by splenic Tr1 cells and functionally moves from the allograft to the spleen.