title: In utero bone marrow transplantation induces donor-specific tolerance by a combination of clonal deletion and clonal anergy.
abstract: OBJECTIVE
  In utero bone marrow transplantation can induce donor-specific tolerance to postnatal solid organ transplantation, although the mechanisms remain poorly defined. In this study, we investigated the role of clonal deletion and clonal anergy in the maintenance of tolerance in a murine model of in utero bone marrow transplantation.
  
  
  METHODS
  DBA/2 mice (MIs(a+)) were used as donors of adult bone marrow, and 14-day-gestation fetal Balb/c mice (MIs(a-)) were used as recipients. Tolerance was defined by donor-specific skin graft survival for more than 8 weeks. Clonal deletion was assessed by flow cytometry for Vbeta6 T cell receptor usage. A tolerant animal demonstrating partial deletion of CD4+/Vbeta6+ T cells and a nontolerant animal were selected for analysis of clonal anergy by a proliferation assay using plate-bound anti-Vbeta6 antibody for stimulation with or without exogenous interleukin-2 (IL2).
  
  
  RESULTS
  Vbeta6+ splenocytes constituted 6.32% of CD4+ T cells in the tolerant animal compared with 9.19% in the nontolerant animal, demonstrating incomplete clonal deletion in the tolerant animal. Stimulation with plate-bound anti-Vbeta6 induced a good proliferative response in the nontolerant animal but a significantly attenuated response in the tolerant animal (P< .001), which was abrogated by the addition of IL2.
  
  
  CONCLUSIONS
  In this murine model of in utero bone marrow transplantation, the tolerant state is characterized by partial clonal deletion of donor reactive T cells and clonal anergy of nondeleted donor reactive T cells. The anergic state can be abrogated by exogenous IL2, suggesting that the mechanism of anergy is a deficiency of IL2 production.
authors.affiliation: The Children's Institute for Surgical Science, The Children's Hospital of Philadelphia, PA 19104, USA.
authors.forename: H B
authors.initials: HB
authors.lastname: Kim
authors.forename: A F
authors.initials: AF
authors.lastname: Shaaban
authors.forename: R
authors.initials: R
authors.lastname: Milner
authors.forename: C
authors.initials: C
authors.lastname: Fichter
authors.forename: A W
authors.initials: AW
authors.lastname: Flake
mesh_terms: D000818:Animals; D016026:Bone Marrow Transplantation / Q000276:immunology*; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D017635:Clonal Anergy; D017637:Clonal Deletion; D005333:Fetus / Q000601:surgery*; D005434:Flow Cytometry; D018655:Lymphocyte Count; D051379:Mice; D008807:Mice, Inbred BALB C; D008811:Mice, Inbred DBA; D013154:Spleen / Q000166:cytology