title: In-vitro generation and characterisation of murine CD4+CD25+ regulatory T cells with indirect allospecificity.
abstract: Naturally arising CD4(+)CD25(+) regulatory T cells play a pivotal role in the prevention of autoimmunity and in the induction of donor-specific transplantation tolerance. Harnessing regulatory cells for potential adoptive cell therapy is hampered by their lack of antigen-specificity and their limited numbers. Here we describe the generation and expansion of murine CD4(+)CD25(+) T cells with antigen-specificity for an K(d) peptide as potential reagents for adoptive cell therapy in promoting donor-specific transplantation tolerance. Using bone marrow-derived autologous dendritic cells pulsed with the K(d) peptide, we generated T cell lines from purified CD4(+)CD25(+) T cells from C56BL/6 mice. The T cell lines expressed high level of CD25 and low level of CD45RB and CD69. They maintained the expression of CD62L, GITR, CTLA-4 and more importantly FoxP3. The CD4(+)CD25(+) T cell lines were anergic after TCR stimulation and produced little cytokine such as IL-2 and IFN-gamma. Importantly, they were more potent than freshly isolated CD4(+)CD25(+) T cells in suppressing proliferation and cytokine secretion by effector CD4(+) T cells. Furthermore, the CD4(+)CD25(+) T cell lines could be expanded to large cell numbers and maintained in culture up to 1 year. The K(d)-specific CD4(+)CD25(+) T cell lines will be invaluable in devising a strategy for the induction of cardiac transplantation tolerance in wild-type B6 mice carrying a full mismatch BALB/c heart.
authors.affiliation: Department of Nephrology and Transplantation, King's College London, Guy's Hospital, London SE1 9RT, United Kingdom.
authors.forename: Julia
authors.initials: J
authors.lastname: Tsang
authors.forename: Shuiping
authors.initials: S
authors.lastname: Jiang
authors.forename: Yakup
authors.initials: Y
authors.lastname: Tanriver
authors.forename: Eva
authors.initials: E
authors.lastname: Leung
authors.forename: Giovanna
authors.initials: G
authors.lastname: Lombardi
authors.forename: Robert I
authors.initials: RI
authors.lastname: Lechler
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology / Q000494:pharmacology; D015703:Antigens, CD / Q000378:metabolism; D000943:Antigens, Differentiation / Q000378:metabolism; D017252:CD3 Complex / Q000276:immunology; D015496:CD4-Positive T-Lymphocytes / Q000166:cytology / Q000276:immunology* / Q000378:metabolism; D060908:CTLA-4 Antigen; D018929:Cell Culture Techniques / Q000379:methods; D002460:Cell Line; D018920:Coculture Techniques; D016207:Cytokines / Q000378:metabolism; D003713:Dendritic Cells / Q000276:immunology; D005434:Flow Cytometry; D051858:Forkhead Transcription Factors / Q000378:metabolism; D053450:Glucocorticoid-Induced TNFR-Related Protein; D006183:H-2 Antigens / Q000276:immunology; D007376:Interleukin-2 / Q000494:pharmacology; D053645:Interleukin-2 Receptor alpha Subunit / Q000276:immunology*; D007519:Isoantigens / Q000276:immunology*; D019041:L-Selectin / Q000378:metabolism; D017493:Leukocyte Common Antigens / Q000378:metabolism; D008213:Lymphocyte Activation / Q000187:drug effects / Q000276:immunology; D051379:Mice; D008810:Mice, Inbred C57BL; D010446:Peptide Fragments / Q000276:immunology; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000276:immunology; D017475:Receptors, Nerve Growth Factor / Q000378:metabolism; D018124:Receptors, Tumor Necrosis Factor / Q000378:metabolism; D050378:T-Lymphocytes, Regulatory / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism
chemical_list: D000906:Antibodies; D015703:Antigens, CD; D000943:Antigens, Differentiation; D017252:CD3 Complex; D060908:CTLA-4 Antigen; C556707:Ctla4 protein, mouse; D016207:Cytokines; D051858:Forkhead Transcription Factors; C498833:Foxp3 protein, mouse; D053450:Glucocorticoid-Induced TNFR-Related Protein; D006183:H-2 Antigens; D007376:Interleukin-2; D053645:Interleukin-2 Receptor alpha Subunit; D007519:Isoantigens; D010446:Peptide Fragments; D016693:Receptors, Antigen, T-Cell, alpha-beta; D017475:Receptors, Nerve Growth Factor; D018124:Receptors, Tumor Necrosis Factor; C106544:Tnfrsf18 protein, mouse; D019041:L-Selectin; D017493:Leukocyte Common Antigens