ACTIVATION AND ROLE OF T REGULATORY CELLS

G. Kinnear¹, K.J. Wood², D. Marshall³, N.D. Jones^1
1Transplantation Research Immunology Group, Nuffield Department of Surgery, Oxford/UNITED KINGDOM, ²Transplantation Research Immunology Group, Nuffield Department of Surgery, University of Oxford, Oxford/UNITED KINGDOM, ³, UCB-Celltech, Slough/UNITED KINGDOM

Body: Introduction: OX40 (CD134) is a member of the tumour necrosis receptor superfamily and is a potent costimulatory molecule that facilitates effector T cell differentiation and survival. Although the impact of blockade of the OX40-OX40L pathway has been well documented in models of autoimmune disease, its effect on the rejection of allografts is less well defined. Methods: The alloimmune response of BM3, alloreactive TCR-transgenic T cells was measured in vitro by the incorporation of ³H thymidine and flow cytometry. In vivo, 1x10⁵ naïve H2K^b-reactive BM3 CD8⁺ T cells were adoptively transferred into syngeneic RAG^-/- mice and the following day mice received an allogeneic H2^b+ skin transplant with and without OX40 blockade. Skin allograft survival was monitored and the BM3 T cell number, CD44 expression (as a marker of antigen experience) and CFSE division profile was determined in the spleen and axillary lymph nodes using flow cytometry. Results: In vitro allogeneic stimulation of BM3 and polyclonal CD8⁺ T cells resulted in the induction of OX40 expression from day 3 of culture. Furthermore, blockade of OX40-OX40L interactions partially inhibited the proliferation of BM3 T cells in vitro (54% inhibition ± 3.5, p<0.0005). Similarly, in vivo, OX40 blockade prevented skin allograft rejection mediated by naïve BM3 T cells compared to controls. However, when OX40 blockade was stopped all skin allografts were rejected (MST 59 vs. 19.5 days; p<0.0014) indicating that tolerance was not induced in this model. Surprisingly, analysis of the lymph nodes draining skin allografts (DLN), 10 days post transplantation, revealed that OX40 blockade had no effect on the activation (CD44 expression) and proliferation of BM3 CD8⁺ T cells to skin allografts. However, in the absence of OX40-OX40L interactions effector BM3 T cells failed to accumulate in the DLN (BM3 T cell number with 6610 ± 1956 or without 43832 ± 14258 OX40 blockade cell number; p<0.05). Conclusion: Taken together, these data demonstrate that activated CD8⁺ T cells express OX40 and that the blockade of OX40-OX40L interactions attenuates CD8⁺ T cell responses to alloantigen in vitro and in vivo by reducing the survival of effector T cells. Therefore, we propose that blocking the OX40-OX40L interaction would be a worthwhile adjunct to pre-existing tolerance induction strategies and may result in the induction of a more reliable and robust form of operational tolerance to allografts.

Disclosure: All authors have declared no conflicts of interest.