Hematopoietic chimerism after non-myeloablative bone marrow transplantation (nmBMT) has been found beneficial for establishment of transplant tolerance. Since bone marrow cells (BMCs) are endowed with “veto” activity, reduction in the number of transferred BMCs usually results in engraftment failure in nmBMT. We previously reported that BM engraftment could be promoted by (1) enhancing Treg activity via iNKT cell stimulation with a liposomal α-galactosylceramide (lipo-aGC), and (2) inducing donor-specific peripheral deletion with CD40/CD40L blockade (MR1). Here, we implemented a 3^rd strategy: additional transfer of donor-derived T cells as veto cells to facilitate BM engraftment. 3 Gy-irradiated BALB/c (H2^d) recipient mice were administered B6 (H2^b) BMCs (5 [times] 10⁶) followed by lipo-aGC plus MR1. On the following day, skin grafts from B6 and C3H (H2^k) mice were simultaneously transplanted on the recipient mice. When this clinically practical number of BMCs was used, all mice rejected both B6 and C3H grafts. However, when 5 [times] 10⁵ splenic T cells obtained from an H2^b+GFP-Tg donor were additionally administered, complete chimerism was established in the mice (fig 1) and they accepted the skin allograft permanently in an H2^b-specific manner. Only H2^b+CD8⁺T cells showed this veto effect, whereas transfer of H2^b+CD4⁺T cells or 3^rd party T cells did not facilitate BM engraftment. Concordantly, early expansion of GFP⁺CD44^hiCD62L^loeffector CD8⁺T cells was observed in many lymphoid organs, including the BM and thymus. BM-derived H2^b+GFP^–T cells immediately regenerated thereafter and had completely replaced GFP⁺T cells until day 60. Therefore, complete chimeric mice did not show GvHD physically and pathologically. Lack of host reactivity was also confirmed in an in vitro proliferation assay. The mice treated with lipo-aGC or MR1 alone rejected BM and skin grafts despite T cell transfer (fig 2), suggesting that Treg cells, peripheral deletion, and veto T cells act in concert for establishing tolerance in nmBMT recipients.

CITATION INFORMATION: Ishii R, Hirai T, Ishii Y, Tanabe K. Combination of Veto Cell Transfer and iNKT Cell Therapy Establishes Complete Hematopoietic Chimerism in Non-Myeloablative BMT Recipients. Am J Transplant. 2016;16 (suppl 3).

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