Purpose: Targeting the process of central (thymic) selection of developing T lymphocytes is the key tolerogenic mechanism of bone marrow transplantation (BM-Tx)-based protocols for transplant tolerance induction. However, they are not amenable to most transplant recipients. Thymic Epithelial Cells (TEC), a population of stromal cells residing in the thymus, exert a major contribution to central selection. However, donor TEC do not develop following BM-Tx protocols. Therefore, we propose a new immunomodulatory strategy based on generating a donor-recipient “Hybrid Thymus”, through donor TEC engraftment into the recipient thymus, to re-engineer the thymic microenvironment that selects developing thymocytes and achieve dominant central tolerance.

Methods: We optimized a protocol for isolating TEC via a combination of negative and positive selection. Purified BALB/c TEC (from 3-12 day old mice) were injected intrathymically into C57BL/6 with or without co-stimulation blockade (CoB: CTLA4-Ig +/- anti-CD40 mAb). TEC survival post-injection was assessed via immunofluorescent staining of thymic sections and peripheral T cells analyzed for changes in TCR Vβ11 expression– a marker of negative selection.

Results: Our optimized purification protocol yields an average 70% TEC purity. Injection of BALB/c TEC in otherwise unmanipulated animals resulted in minimal survival by POD21, suggesting an absence of intrinsic immunomodulatory capacity of these cells. However, CTLA4-Ig co-administration exerted a significant protection with a higher survival at POD21 and 28. In this latter treatment group, the percentage of peripheral Vβ11+ T cells on d21 was significantly decreased indicating actuation of negative selection by the engrafted donor TEC. The full CoB regimen provided the highest survival rate at all time points, indicating the need for a complementary treatment to promote alloTEC engraftment. We are currently investigating the long term TEC survival and the impact on host alloreactivity.

Conclusion: Our preliminary data show that the thymic engraftment, survival, and function of allogenic TEC can be promoted by CoB. These exciting results indicate that engineering a donor-recipient Hybrid Thymus is feasible and has the potential to promote a dominant regulation of alloreactivity that could be conducive to transplant tolerance induction.

CITATION INFORMATION: AlFadil S, Kim M.-J, Iglesias Lozano M, Oh B, Lee W, Brandacher G, Serworld T, Raimondi G. Engineering a “Hybrid Thymus” to Promote Transplant Tolerance. Am J Transplant. 2016;16 (suppl 3).

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