Purpose: Peri-operative cadaveric donor vertebral body bone marrow (VBBM) infusion has shown promising immunomodulatory effects (graft maintenance on low-dose tacrolimus monotherapy) in clinical trials of hand transplantation. Mechanisms behind such effects are not yet well-characterized.
Methods: VBBM was prepared, cryopreserved, and thawed as in clinical trials. Samples were stained with monoclonal antibodies to detect various cell populations including subsets of T lymphocyte and mesenchymal stem cell (MSC) populations. Antibodies included anti-human CD3, CD4, CD8, CD45RA, CD34, CD25, Foxp3, CD127, TCRΑΒ and markers for Th1, Th2, and Th17 cells. MSCs were detected as CD34–/11b–/19–/45–/HLA-DR–/CD73+CD105+CD90+ or CD271+cells. Conventional iliac-crest bone marrow aspirates from consenting donors were similarly analyzed. Mixed lymphocyte reaction assays: allogeneically stimulated CFSE-stained PBMCs were co-cultured with whole bone marrow or flow cytometrically sorted bone marrow subpopulations as regulators for 7 days. Results were analyzed by flow cytometry.
Results: VBBM contains cell populations with known immunomodulatory properties. Composition is grossly comparable to conventionally obtained bone marrow, with differences in some populations (partial list shown).
Both whole bone marrow and sorted sub-populations demonstrated immunomodulatory properties. Whole bone marrow co-cultured in a 5:1 responder-regulator ratio suppressed allogeneic stimulation by 90.8%. The T regulatory cell (CD4+CD25highCD127lo) population was particularly potent, suppressing allogeneic stimulation by 95.4% when cultured in a 3000:1 responder-regulator ratio.
Conclusion: VBBM contains cell populations with potent immunomodulatory effects. Upcoming studies include cytokine analysis and donor-specific vs. third-party experiments. Our ongoing immunophenotypical and functional characterization helps identify the mechanism behind VBBMs immunomodulatory properties, paving the way towards opportunities to refine the cell product and maximize its clinical effectiveness.
To cite this abstract in AMA style:Yuan N, Sarkar K, Lemas M, Bitzan J, Sproul J, Lee W, Brandacher G, Cooney D. In Vitro Characterization of Vertebral Bone Marrow Used for Immunomodulation in Hand Transplantation [abstract]. Am J Transplant. 2013; 13 (suppl 5). https://atcmeetingabstracts.com/abstract/in-vitro-characterization-of-vertebral-bone-marrow-used-for-immunomodulation-in-hand-transplantation/. Accessed August 20, 2019.
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