Background: Myeloid-derived suppressor cells (MDSC) have shown promise in inhibiting allograft rejection and in promoting transplant tolerance in rodent models. We recently reported that monocytic (m)MDSC isolated from cytokine-mobilized (rhGM-CSF, rhG-CSF) rhesus macaque PBMC could markedly suppress proliferation of CD3/CD28-stimulated autologous T cells and enhance Treg. Given the very limited number of mMDSC that could be flow-sorted from mobilized PBMC for adoptive transfer, we examined whether mMDSC could be generated from rhesus bone marrow (BM) cells.

Methods: Femural BM cells (20×10⁶)were cultured in T75cm² flasks for 7d in DMEM media supplemented with 7% v/v human AB serum, rhGM-CSF (80ng/ml) and rhIL-6 (15ng/ml). FACS was used to isolate putative mMDSC that were HLA-DR^–, CD3^–, CD20^–, CD33⁺, CD14^hi+, CD34^– and CD11b⁺. The ability of these putative MDSC to suppress proliferation of autologous T cells was assessed by co-culture of flow-sorted 50,000 (1:2) or 25,000 (1:4) MDSC per 100,000 T cells for 4d in the presence of anti-CD3/CD28 beads (1 bead : 15 T cells). Autologous T cells incubated in media alone or with anti-CD3/CD28 beads were included as controls.

Results The mean incidence of mMDSC that could be identified in 7d BM cell cultures stimulated with GM-CSF and IL-6 was 7.3±1.3 %. At a ratio of 1 mMDSC: 2 CD3⁺ T cells, these MDSC suppressed CD4⁺ T cell proliferation by a mean of 90% and CD8⁺ T cell proliferation by 70%. This suppressive activity of BM-derived mMDSC was more potent (on a per cell basis) than that of mMDSC sorted from mobilized PBMC. Also, the BM-derived mMDSC increased the incidence of CD4⁺CD25⁺CD127^–Foxp3⁺ Treg more effectively than mMDSC for mobilized PBMC.

Conclusions: These results show that monocytic MDSC can be generated from BM of healthy (non-mobilized) rhesus monkeys, enhancing the numbers of these regulatory myeloid cells available for flow sorting and adoptive cell transfer to allograft recipients. These BM-derived mMDSC are more potent suppressors of T proliferation than those isolated from cytokine-mobilized donor PBMC and promote increased numbers of Treg relative to T effector cells.

CITATION INFORMATION: Thomson A, Zahorchak A, Perez-Gutierrez A, Ezzelarab M. Generation of Potent Monocytic Myeloid-Derived Suppressor Cells from Rhesus Macaque Bone Marrow. Am J Transplant. 2017;17 (suppl 3).

« Back to 2017 American Transplant Congress