IntroductionIn the peripheral lymphoid tissues of normal mice and humans, 1% to 5% of ␣ T-cell receptor-positive (TCR ϩ ) T cells are CD4 Ϫ CD8 Ϫ (double-negative [DN]) T cells. 1,2 MRL/Mpj-lpr/lpr mice have a mutant Fas gene and a massive lymphadenopathy consisting of an age-related accumulation of DN ␣TCR ϩ T cells. 3,4 The DN T cells have been shown to possess the capacity to regulate auto-and alloimmune responses and induce immune tolerance. [5][6][7][8] However, the origin of peripheral DN T cells is still unclear. The heterogeneity of DN T-ells in the expression of surface markers suggests that several maturation/differentiation pathways may exist. In murine models, several studies have demonstrated that DN ␣ TCR ϩ T cells can be derived directly from CD8 ϩ T cells. [9][10][11][12] Other studies suggest that DN ␣ TCR ϩ natural killer T cells (NKT cells) arise extrathymically from bone marrow (BM). 13 More recently, Ford et al reported that DN T regulatory cells can develop outside the thymus, but not from mature CD8 ϩ T-cell precursors. 14 However, a differentiation pathway of peripheral DN T cells from CD4 ϩ T cells was not identified.In this report, we monitored CD4 expression during CD4 ϩ T-cell proliferation and differentiation and identified a new pathway for the generation of a DN regulatory T-cell subset. This pathway uncovered a new intrinsic homeostatic mechanism that regulates the magnitude of immune responses to alloantigen both in vitro and in vivo. Our observations will permit the development of novel, cell-based, therapeutic approaches for the prevention of allograft rejection and for the treatment of autoimmune diseases. Materials and methodsMice Male C57BL/6 (H-2 b ), C57BL/6 congenic for CD45.1, C57BL/6 TEa TCR-transgenic, C57BL/6 perforin gene knock-out (KO), C57BL/6 RAG Ϫ/Ϫ , DBA/2 (H-2 d ), C3H (H-2 k ), and B6D2F1 (H-2 b/d ) mice were obtained from The Jackson Laboratory (Bar Harbor, ME). Foxp3 gfp knock-in C57BL/6 mice were provided by Dr Wenda Gao (Boston, MA). 15 All mice were maintained in the animal facilities of Harvard Institutes of Medicine. Reagents and antibodiesRecombinant mouse interleukin-2 (IL-2), IL-4, and granulocyte-macrophage colony-stimulating factor (GM-CSF) were obtained from Biosource (Camarillo, CA). CD4 ϩ T-cell enrichment column, T-cell enrichment column, and recombinant mouse IL-15 were obtained from R&D Systems (Minneapolis, MN). Fluorochrome-conjugated antibodies to mouse CD3, CD4, CD8, CD25, CD28, CD40, CD44, CD45.1, CD69, CD86, Ter119, B220, CD11b, CD11c, Gr1, NK1.1, TCR, TCR␥␦, and isotype controls were obtained from eBioscience (San Diego, CA). Annexin V-PE was purchased from BD Pharmingen (San Diego, CA). CD4 ϩ CD25 ϩ regulatory T cell (Treg) isolation kits, anti-PE microbeads, and magnetic bead separation columns were obtained from Miltenyi Biotec (Auburn, CA). Mitomycin C was obtained from Sigma (St Louis, MO).Purification of CD4 ؉ , CD4 ؉ CD25 ؉ , CD4 ؉ CD25 ؊ , and CD4 ؊ CD8 ؊ DN T cellsSingle-cell suspensions were prepared from the spleens and...
Preserving and enhancing the primary function of transplanted islets is not only crucial for improving the outcome of the islet transplantation, but is also important for reducing the islet mass required to achieve insulin independence. Uncoupling protein 2 (UCP2) is a member of the uncoupling protein family, which is localized to the inner mitochondrial membrane and negatively regulates insulin secretion in the pancreatic β-cells. In this study, we assessed the importance of UCP2 in improving islet graft primary function by using UCP2 gene-knockout (UCP2-KO) mice in a syngeneic islet transplantation model. Islets were isolated from UCP2-KO or wild-type (WT) C57BL/6J mice. The effects of deficiency of UCP2 on islet transplantation and islet function were determined. Two hundred islets from UCP2-KO, but not from WT, donors were capable of completely restoring normoglycemia in 1 week in all syngeneic diabetic recipients. Islets harvested from UCP2-KO mice secreted onefold more insulin in GSIS assay than that from WT mice, and maintained normal GSIS after 72-h exposure to high glucose challenge. In addition, UCP2-KO islets expressed twohold higher Bcl-2 mRNA than that from WT islets, and were resistant to high glucose and proinflammatory cytokine induced death. Our study explored a potential mechanism that may explain the benefit of UCP2-KO islets in islet transplantation. Targeting UCP2 may provide a novel strategy to improve primary function of transplanted islets and reduce the number of islets required in transplantation.Key words: Uncoupling protein 2 (UCP2); Diabetes; Islets transplant; Primary function INTRODUCTIONTransplanted islets are particularly vulnerable in the immediate posttransplantation period (7). Recent study indicated recipient hyperglycemia rendered islet grafts Diabetes mellitus is now fast emerging as one of the biggest health catastrophes in modern society. It was essusceptible to dysfunction and failure. An increased incidence of primary nonfunction was observed when a timated that approximately 160 million people worldwide suffered from diabetes in 2000, and this number marginal number of islets were transplanted into severely diabetic mice, in comparison with moderately diwas projected to increase to 221 million in 2010 and to 366 million in 2030 (25). abetic mice (12). Hyperglycemia increased oxidative stress and deteriorated β-cell function in transplanted isRecent progress in the pancreas' enzymatic digestion process along with novel immunosuppression strategies lets. Islet graft response to transplantation injury includes upregulation of protective as well as apoptotic has led to successful clinical trials of islet transplantation in humans (18). However, successful islet transplangenes (17). Therefore, preserving and enhancing the primary function of transplanted islets are not only crucial tation depends on the infusion of higher mean islet mass (>10,000 IE/kg) prepared from 2-4 donor pancreases to for improving the outcome of the islet transplantation, but also important for reduc...
CD4−CD8− (double-negative, DN) T-cells are capable of down-regulating the immune response. However, the origin and developmental pathway of DN T-cells are still unclear. In this study, we showed that proliferated CD4+ T-cells could convert to DN T-cells after 4–5 rounds of either allogeneic or syngeneic DC triggered proliferation, IL-2 and IL-15 enhanced the conversion. The disappearance of CD4 molecular on converted DN T-cells was a result of CD4 gene silence. Converted DN T-cells were resistant to AICD and expressed unique cell markers and gene profile. Moreover, DN T-cells retained a stable phenotype after re-stimulation. DN T-cells converted from CD4+ T-cells exerted powerful inhibition on same, but not third party, alloantigen triggered proliferation of naïve T cells. Perforin, a cytotoxic lymphocyte related cytokine, was highly expressed by DN T-cells, played a role in DN T-cell mediated suppression. Transferring of DN T-cells alone could prevent allograft rejection and the onset of autoimmune disease in vivo. This previously unknown pathway of differentiating DN regulatory T-cells represents a negative feedback mechanism of immune response. The results of using ex vivo CD4+ T-cells converted DN T-cells in transplantation and autoimmune disease models support the concept and the feasibility of potentially utilizing this novel cell-based therapy.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.