title: CD27/CFSE-based ex vivo selection of highly suppressive alloantigen-specific human regulatory T cells.
abstract: Naturally occurring CD4(+)CD25(+) regulatory T cells (Treg) are crucial in immunoregulation and have great therapeutic potential for immunotherapy in the prevention of transplant rejection, allergy, and autoimmune diseases. The efficacy of Treg-based immunotherapy critically depends on the Ag specificity of the regulatory T cells. Moreover, the use of Ag-specific Treg as opposed to polyclonal expanded Treg will reduce the total number of Treg necessary for therapy. Hence, it is crucial to develop ex vivo selection procedures that allow selection and expansion of highly potent, Ag-specific Treg. In this study we describe an ex vivo CFSE cell sorter-based isolation method for human alloantigen-specific Treg. To this end, freshly isolated CD4(+)CD25(+) Treg were labeled with CFSE and stimulated with (target) alloantigen and IL-2 plus IL-15 in short-term cultures. The alloantigen-reactive dividing Treg were characterized by low CFSE content and could be subdivided by virtue of CD27 expression. CD27/CFSE cell sorter-based selection of CD27(+) and CD27(-) cells resulted in two highly suppressive Ag-specific Treg subsets. Each subset suppressed naive and Ag-experienced memory T cells, and importantly, CD27(+) Treg also suppressed ongoing T cell responses. Summarizing, the described procedure enables induction, expansion, and especially selection of highly suppressive, Ag-specific Treg subsets, which are crucial in Ag-specific, Treg-based immunotherapy.
authors.affiliation: Department of Blood Transfusion and Transplantation Immunology, Radboud University Nijmegen Medical Center, Nijmegen, The Netherlands. h.koenen@abti.umcn.nl
authors.forename: Hans J P M
authors.initials: HJ
authors.lastname: Koenen
authors.forename: Esther
authors.initials: E
authors.lastname: Fasse
authors.forename: Irma
authors.initials: I
authors.lastname: Joosten
mesh_terms: D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D002450:Cell Communication / Q000276:immunology; D002454:Cell Differentiation / Q000276:immunology*; D002469:Cell Separation / Q000379:methods*; D002478:Cells, Cultured; D018920:Coculture Techniques; D018984:Epitopes, T-Lymphocyte / Q000276:immunology*; D005452:Fluoresceins / Q000032:analysis*; D005456:Fluorescent Dyes / Q000032:analysis*; D006801:Humans; D007156:Immunologic Memory / Q000276:immunology; D007519:Isoantigens / Q000276:immunology*; D008213:Lymphocyte Activation / Q000276:immunology; D015375:Receptors, Interleukin-2 / Q000378:metabolism; D015398:Signal Transduction / Q000276:immunology; D013388:Succinimides / Q000032:analysis*; D050378:T-Lymphocytes, Regulatory / Q000166:cytology / Q000276:immunology* / Q000378:metabolism; D018127:Tumor Necrosis Factor Receptor Superfamily, Member 7 / Q000032:analysis* / Q000096:biosynthesis / Q000502:physiology
chemical_list: C087165:5-(6)-carboxyfluorescein diacetate succinimidyl ester; D018984:Epitopes, T-Lymphocyte; D005452:Fluoresceins; D005456:Fluorescent Dyes; D007519:Isoantigens; D015375:Receptors, Interleukin-2; D013388:Succinimides; D018127:Tumor Necrosis Factor Receptor Superfamily, Member 7