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Comparative analysis of bone marrow versus blood monocyte-derived regulatory dendritic cells for evaluation in a miniature swine vascular composite allotransplantation model
Tarek Elgendy1, Deokyeol Kim2, Alan Zahorchak1, Mohamed Ezzelarab1, Wensheng Zhang2, Marta Minervini3, Kia Washington2, Mario Solari2 and Angus W. Thomson1
1Starzl Transplantation Institute, Department of Surgery, 2Department of Plastic Surgery, 3Departmen
2018-01-29
Presenter: Tarek Elgendy
Affidavit:
Large animal studies are a team effort. Dr. Elgendy is the team leader on this project and his contributions meet my standards for 1st authorship.
Director Name: MGS
Author Category: Fellow Plastic Surgery
Presentation Category: Basic Science Research
Abstract Category: General Reconstruction
Background: Dendritic cell therapy is a promising approach to reduce dependence on immunosuppressive drugs and to promote transplant tolerance. Our aim was to characterize for the first time miniature swine regulatory dendritic cells (DCreg) generated from either blood monocytes or bone marrow (BM) cells for evaluation in a pre-clinical vascular composite allograft model.
Methods: Blood or femoral BM was obtained from class I and class II MHC-defined miniature swine. Control immature DC (imDC) were generated from either CD14+ bead-isolated blood monocytes or BM cells over 7 days culture in media supplemented with 10% FCS, porcine (p)GM-CSF and pIL-4. To generate DCreg, vitamin D3 was added on day 0, and VitD3 and pIL-10 (60 ng/ml) added on day 4. Cell surface phenotype was determined by staining the DC with fluorochrome-conjugated antibodies against SLA-DR, CD80/CD86, CD172a, CD1 and CD3 and by comparing mean fluorescence intensity and percent positive cells. The ability of the DCreg to stimulate allogeneic T cell proliferation was determined by carboxyfluorescein succinimidyl ester mixed leukocyte reaction.
Results: Swine BM-derived DCreg were poor allogeneic T cell stimulators compared to mature DC. After LPS challenge (DCregLPS), levels of T cell co-stimulatory molecules remained low and the DCreg remained poor allostimulators. Similar results were obtained from blood. However, adequate numbers of DCreg for prospective adoptive cell therapy could be generated more readily from BM.
Conclusions: Swine DCreg that resist maturation in response to a potent inflammatory stimulus can be generated efficiently from BM to achieve adequate numbers for adoptive cell therapy.
