An irradiation-free nonmyeloablative bone marrow transplantation model: Importance of the balance between donor T-cell number and the intensity of conditioning

Background. Animal allogeneic bone marrow transplantation (BMT) models with nonmyeloablative conditioning regimens have so far required irradiation or antibodies in addition to immunosuppressive drugs for engraftment. Moreover, although it is known that the balance between donor T-cell number and the dose of immunosuppressive drugs would be critical for engraftment, it has not been experimentally clarified in a nonmyeloablative regimen. Methods. We used C57BL/6 mice as donors and DBA/2 mice as recipients with a nonmyeloablative regimen including fludarabine (Flu) and cyclophosphamide (CPA) without irradiation or antibodies. To determine the adequate doses, we injected recipients with various doses of Flu and CPA, and 2 × 10⁷ bone marrow cells (BMC) and 5 × 10⁷ splenocytes (SC). Furthermore, using T-cell-depleted BMC and enriched T cells, we investigated the balance between donor T-cell number and the dose of Flu. Results. Doses of Flu at 150 mg/kg/dayX6 and CPA at 150 mg/kg/dayX2 were most appropriate for engraftment with low mortality. All mice appropriately pretreated and transplanted with both BMC and SC exhibited complete donor chimeras. Donor cell engraftment was not enhanced by any increase of BMC transplanted, and dose escalation of donor T cells but not BMC led to the reduction of Flu dose required for engraftment of donor cells. Conclusions. We have established a murine nonmyeloablative BMT model in a fully MHC-mismatched combination for donor cell engraftment with complete donor chimerism. Simultaneously, we have quantitatively demonstrated that the balance between donor T-cell number and the dose of immunosuppressive drugs is critical for stable engraftment.