title: A converse 4-1BB and CD40 ligand expression pattern delineates activated regulatory T cells (Treg) and conventional T cells enabling direct isolation of alloantigen-reactive natural Foxp3+ Treg.
abstract: Natural regulatory T cells (nTreg) play a central role in the induction and maintenance of immunological tolerance. Experimental transplant models and recent clinical trials demonstrate that nTreg can control alloreactivity. To upgrade Treg-based cell therapies to a selective suppression of undesired immune reactions, only the transfer of Ag-specific nTreg represents the appropriate therapeutic option. However, Ag-specific nTreg are present at extremely low frequencies in the periphery, and so far appropriate surface markers for their precise identification are missing. In this study, we demonstrate that activated nTreg and activated conventional T cells differ in their 4-1BB and CD40 ligand (CD40L) expression signatures, allowing a clear dissection from each other. Based on the expression of 4-1BB and absence of CD40L expression, human alloantigen-reactive Foxp3(+) nTreg can be directly isolated from MLR cultures with high purity. Alloantigen-reactive 4-1BB(+)CD40L(-) nTreg were characterized by a completely demethylated Treg-specific demethylated region and showed alloantigen-specific suppressive properties superior to polyclonal Treg. Importantly, isolated 4-1BB(+)CD40L(-) nTreg maintain the nTreg phenotype and alloantigen-reactivity after in vitro expansion. Our results offer the possibility to simultaneously analyze Ag-specific nTreg and conventional T cells, and to establish cellular therapies with Ag-specific nTreg aiming at a specific inhibition of unwanted immunity.
authors.affiliation: Regenerative Immunology and Aging, Berlin-Brandenburg Center for Regenerative Therapies, Charité University Medicine, 13353 Berlin, Germany.
authors.forename: Anne
authors.initials: A
authors.lastname: Schoenbrunn
authors.forename: Marco
authors.initials: M
authors.lastname: Frentsch
authors.forename: Siegfried
authors.initials: S
authors.lastname: Kohler
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Keye
authors.forename: Hans
authors.initials: H
authors.lastname: Dooms
authors.forename: Beate
authors.initials: B
authors.lastname: Moewes
authors.forename: Jun
authors.initials: J
authors.lastname: Dong
authors.forename: Christoph
authors.initials: C
authors.lastname: Loddenkemper
authors.forename: Joachim
authors.initials: J
authors.lastname: Sieper
authors.forename: Peihua
authors.initials: P
authors.lastname: Wu
authors.forename: Chiara
authors.initials: C
authors.lastname: Romagnani
authors.forename: Nadine
authors.initials: N
authors.lastname: Matzmohr
authors.forename: Andreas
authors.initials: A
authors.lastname: Thiel
mesh_terms: D053330:4-1BB Ligand / Q000096:biosynthesis* / Q000235:genetics*; D000818:Animals; D023201:CD40 Ligand / Q000096:biosynthesis* / Q000172:deficiency / Q000235:genetics*; D002469:Cell Separation; D002478:Cells, Cultured; D004195:Disease Models, Animal; D051858:Forkhead Transcription Factors / Q000096:biosynthesis*; D006086:Graft vs Host Disease / Q000276:immunology; D006801:Humans; D007519:Isoantigens / Q000276:immunology* / Q000378:metabolism; D008213:Lymphocyte Activation / Q000235:genetics / Q000276:immunology*; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D016176:T-Lymphocyte Subsets / Q000276:immunology / Q000378:metabolism; D050378:T-Lymphocytes, Regulatory / Q000276:immunology* / Q000378:metabolism / Q000637:transplantation; D014183:Transplantation, Heterologous
chemical_list: D053330:4-1BB Ligand; C418974:FOXP3 protein, human; D051858:Forkhead Transcription Factors; D007519:Isoantigens; D023201:CD40 Ligand