title: Requirements for prolongation of allograft survival with regulatory T cell infusion in lymphosufficient hosts.
abstract: BACKGROUND
  For the clinical applicability of regulatory T cells (Tregs) in transplantation, it is critical to determine if donor antigen specificity is required for their immunosuppressive function. We developed an allospecific CD4(+) T cell receptor transgenic (TCR-tg) mouse as a source for large numbers of Tregs with defined allospecificity and tested whether they are more effective than polyclonal Tregs at suppressing allograft rejection.
  
  
  METHODS
  CD4(+)CD25(+)CD62L(hi) T cells were sorted from the spleen and peripheral lymph nodes of wild-type (WT-Tregs) and TCR-tg (Allo-Tregs) mice, and expanded using IL-2 and anti-CD3/anti-CD28 conjugated magnetic beads. Tregs were tested for their ability to suppress the proliferation and cytokine production of alloreactive CD4(+)CD25(-) T cells in mixed leukocyte assays. Syngeneic WT hosts were adoptively transferred 5 × 10(6) Tregs and transplanted with allogeneic hearts.
  
  
  RESULTS
  Using anti-CD3/anti-CD28 conjugated beads, Tregs were expanded in vitro 100-fold and maintained their suppressor phenotype and function. Allo-Tregs were 6-8 times more potent on a cell-for-cell basis than WT-Tregs in suppressing allospecific proliferation in vitro. Allo-Tregs were unable to suppress in the absence of allo-antigen. Adoptive transfer of expanded Allo-Tregs into WT recipients prolonged the graft survival in a F1 heart transplant model compared with WT-Treg or no treatment [20.0 ± 4.4 d (n = 6) versus 10.4 ± 1.2 (n = 8) and 9.7 ± 1.6 d (n = 6)].
  
  
  CONCLUSIONS
  Unlike polyclonal Tregs, allospecific Tregs are able to prolong allograft survival. However, large numbers of Allo-Tregs were unable to induce tolerance, suggesting that Treg therapy in immunocompetent recipients will require conditioning and/or additional immunomodulation for the induction of tolerance.
authors.affiliation: Department of Surgery, Duke University Medical Center, Durham, North Carolina 27710, USA. todd.brennan@duke.edu
authors.forename: Todd V
authors.initials: TV
authors.lastname: Brennan
authors.forename: Qizhi
authors.initials: Q
authors.lastname: Tang
authors.forename: Feng-Chun
authors.initials: FC
authors.lastname: Liu
authors.forename: Vunghi
authors.initials: V
authors.lastname: Hoang
authors.forename: Mingying
authors.initials: M
authors.lastname: Bi
authors.forename: Jeffrey A
authors.initials: JA
authors.lastname: Bluestone
authors.forename: Sang-Mo
authors.initials: SM
authors.lastname: Kang
mesh_terms: D000818:Animals; D049109:Cell Proliferation; D006084:Graft Rejection / Q000276:immunology; D006085:Graft Survival / Q000276:immunology*; D016027:Heart Transplantation / Q000276:immunology*; D007121:Immunocompetence / Q000276:immunology*; D056747:Immunomodulation; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D023421:Models, Animal; D050378:T-Lymphocytes, Regulatory / Q000276:immunology* / Q000637:transplantation*; D023001:Transplantation Tolerance / Q000276:immunology; D014184:Transplantation, Homologous