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Abstract

Aim Stable chimerism and tolerance can be achieved in HLA-disparate kidney recipients treated with donor hematopoietic stem cell (DHCS) transplantation supplemented with facilitating cells (FC) derived from mobilized peripheral blood (mPB) after living donor leukopheresis (Sci. Transl. Med. 2012; 4 (124): 124re28). The use of deceased donor (dd) vertebral body bone marrow (VBM), however, could extend the protocol to multiple dd transplant recipients. We aimed to characterize and evaluate the functional capacity of dd bone marrow-derived FC. Methods VBM was manually minced and gently shaken in RPMI ± mobilizing agent AMD-3100. Magnetic microbeads and flow sorting were used to purify DHSC and FC based on CD34+ and CD8a+/TCR αβ - expression, respectively (n = 10). Cells were seeded in allogeneic and autologous mixed lymphocyte cell reactions (MLR) to test modulatory effects of DHSC and FC (n = 4). Results Incubation of the VBM tissue in AMD-3100 did not significantly enhance cell yield. VBM FC yield was similar to mPB yield (2.05 ± 0.92% (n = 9) vs. 2.06 ± 1.28% (n = 5), respectively. FC cells derived from VBM had similar cell markers as those described in mPB and exhibit a heterogenous cell phenotype, including NK and pre-plasmacytoid dendritic markers CD56 (47.95% ± 0.07), CD3 (31.85% ± 2.33), and CD11b (40.55% ± 8.41). Notably, VBM or mPB-derived FC inhibited proliferation of CFSE-labeled CD4+ responder cells stimulated with allogeneic cells (15% FC vs 26% control in VBM; 0.46% FC vs 17.8% control in mPB) and increased the expression of CD127-CD4+CD25high regulatory T cells. The inhibitory and Treg amplification properties were similar to those of CD34+ cells from VBM and mPB. Conclusions Although the gross make up of the VBM differs from that of mPB, the phenotypic and functional characteristics of FC are similar. These results suggest VBM-derived FC can also be used in transplant recipients for enhancing stable chimerism.