Prolongation of Allograft Survival by Intrathymic Introduction of Allogeneic Thymic Epithelial Cells

Abstract

Thymic epithelial cells (TECs), the primary stromal population in the thymus, are critical to establish the microenvironments required for generating self-tolerant, pathogen-responsive T cells. However, due to the inaccessibility of the thoracic cavity and the fragility of TECs, manipulating the thymus to modulate adaptive immune responses remains challenging. We developed a novel surgical procedure that allowed access to thymus glands for intrathymic injection (ITI), without the need to cut the manubrium of the sternum bone. Horizontal cuts of the muscle layers between the first and second bones of the rib cage expose the thymus with minimal side effects. This novel ITI procedure enabled us to implant 16.5-day embryonic TECs into an adult syngeneic recipients’ thymus. Long-term survival and colonization of the implanted TECs was observed via live imaging, flow cytometry, and immunohistochemistry. We further investigated if introduction of allogeneic TECs in the thymus promote negative selection against allo-reactive T cells and induce donor-specific immune tolerance. We engrafted B6 islets under the kidney capsules of chemically induced diabetic Balb/C mice, preconditioned with T-cell depletion sera and intrathymic injection of B6 16.5-day embryonic TECs. Prolonged survival of the transplanted islets was observed compared to sham-operated controls. Mixed lymphocyte reaction experiments further showed dampened T cell proliferation responses against donor B6 antigens. Moreover, increased levels of Foxp3+ T-regulatory cells were detected in Balb/C recipients with ITI of B6 TECs, suggesting that implanted TECs could also promote the generation of alloantigen-specific T regulatory cells in addition to facilitating negative selection of donor antigen-reactive T cells. Our results suggest that intrathymic transplantation of TECs could be an effective way to induce donor-specific immune tolerance with limited general immune suppression. Disclosure I. Pradhan: None. S. Wong-Noonan: None. S. Bertera: None. R.J. Lakomy: None. H. Cohen: None. D. Wells: None. M. Trucco: None. Y. Fan: None.