Ethylenecarbodiimide-fixed donor splenocyte infusions differentially target direct and indirect pathways of allorecognition for induction of transplant tolerance

J Immunol. 2012 Jul 15;189(2):804-12. doi: 10.4049/jimmunol.1103705. Epub 2012 Jun 13.


Strategic exposure to donor Ags prior to transplantation can be an effective way for inducting donor-specific tolerance in allogeneic recipients. We have recently shown that pretransplant infusion of donor splenocytes treated with the chemical cross-linker ethylenecarbodiimide (ECDI-SPs) induces indefinite islet allograft survival in a full MHC-mismatched model without the need for any immunosuppression. Mechanisms of allograft protection by this strategy remain elusive. In this study, we show that the infused donor ECDI-SPs differentially target T cells with indirect versus direct allospecificities. To target indirect allospecific T cells, ECDI-SPs induce upregulation of negative, but not positive, costimulatory molecules on recipient splenic CD11c(+) dendritic cells phagocytosing the injected ECDI-SPs. Indirect allospecific T cells activated by such CD11c(+) dendritic cells undergo robust initial proliferation followed by rapid clonal depletion. The remaining T cells are sequestered in the spleen without homing to the graft site or the graft draining lymph node. In contrast, direct allospecific T cells interacting with intact donor ECDI-SPs not yet phagocytosed undergo limited proliferation and are subsequently anergized. Furthermore, CD4(+)CD25(+)Foxp3(+) T cells are induced in lymphoid organs and at the graft site by ECDI-SPs. We conclude that donor ECDI-SP infusions target host allogeneic responses via a multitude of mechanisms, including clonal depletion, anergy, and immunoregulation, which act in a synergistic fashion to induce robust transplant tolerance. This simple form of negative vaccination has significant potential for clinical translation in human transplantation.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adoptive Transfer / methods
  • Animals
  • Antigen-Presenting Cells / immunology
  • Antigen-Presenting Cells / metabolism
  • Carbodiimides / administration & dosage*
  • Cross-Linking Reagents / administration & dosage
  • Gene Knock-In Techniques
  • Graft Survival / immunology
  • Infusions, Intravenous
  • Isoantigens / administration & dosage
  • Isoantigens / immunology
  • Isoantigens / metabolism*
  • Male
  • Mice
  • Mice, Inbred BALB C
  • Mice, Inbred C57BL
  • Mice, Knockout
  • Mice, Transgenic
  • Phagocytes / immunology
  • Phagocytes / metabolism
  • Signal Transduction / immunology*
  • Spleen / cytology
  • Spleen / immunology*
  • Spleen / transplantation*
  • Transplantation Tolerance / immunology*


  • Carbodiimides
  • Cross-Linking Reagents
  • Isoantigens