CD8⁺/TCR^– facilitating cells (FCs) are a rare bone marrow-derived cell population that enhances engraftment of hematopoietic stem cells (HSC) in major histocompatibility complex disparate recipients without causing graft versus host disease. Both human and mouse FC exert a number of regulatory effects on HSC, including the enhancement of survival and clonogenicity of HSCs and the induction of regulatory cells including IL-10-producing Tr1 cells, Tregs, and Bregs. In the present study we evaluated the ontogeny of mouse FCs after infusion. FACS sorted 200,000 FCs from C57BL/6 (45.2) mice were transplanted to B6SJL (45.1) mice conditioned with 500cGy. Donor FC-derived cells were analyzed three weeks later in recipient bone marrow, spleen, thymus and lymph nodes (LN). We found that donor FCs do not retain their CD8⁺TCR^– phenotype and were mainly found in spleen and lymph-nodes (LN). The majority of donor cells in spleen and LN were CD19⁺ B cells, whereas in the thymus the pre-dominant phenotype was CD4⁺CD8⁺T cells. This suggests that FCs contain lymphoid progenitor cells that differentiate into either B or T cells depending upon the microenvironment. Donor derived myeloid cells were also found in low frequencies. To better characterize B cell progenitors in mouse FCs, 50,000 CD8⁺TCR^– FCs sorted from 6-8 week oldC57BL/6 mice were stained with an eight-color immunophenotyping panel composed of CD8, TCRαβ, TCRγδ, B220, CD43, CD93, CD24, BP1 and IgM. Among CD8⁺TCR^– total FC (n=8), 58% were pro-B B cell progenitors (B220⁺CD43⁺CD24⁺IgM^–BP1^–), 1.4 % were immature B cell progenitors (B220⁺CD43^–CD24⁺IgM⁺CD93⁺), and only 0.3% were pre-B cell progenitors (B220⁺CD43^–CD24⁺IgM^–BP1⁺).

To exclude the contribution of preferential proliferation of pre-existing CD19⁺ (mature B cells) in the FCs, we transferred sorted CD19-depleted FCs. Depletion of the CD19⁺ expressing cells from FCs did not affect the predominance of CD19⁺ cells in spleen and LNs, supporting that FCs may contain lymphoid progenitors which differentiate into CD19⁺ B cells. Interestingly, a more pre-dominant T cell population was observed at five weeks, and about 5% of CD4⁺T cells expressed Foxp3. Our study shows for the first time that FCs may contain common lymphoid progenitor cells that give rise to B cells, T cells and regulatory T cells in nonmyeloablatively conditioned congenic mice.

CITATION INFORMATION: Chhabra A, Wen Y, Huang Y, Xu H, Merchak A, Ildstad S. Mouse FCs May Contain Lymphoid Progenitor Cells That Have the Capacity to Generate B and T Cells In Vivo. Am J Transplant. 2017;17 (suppl 3).

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