In September of 1994, and subsequently in November of 1996, this center embarked on studies of donor-specific bone marrow cell (DBMC) infusion first in cadaver renal transplant recipients (n=63) and then in recipients of living-related donor kidneys (n=48). The goal was (and continues to be) to eventually withdraw immunosuppression by identifying states of operational immunological tolerance that might be induced by DBMC and continuously monitored. Two significant observations have emerged: 1) There is a positive correlation of DBMC chimerism with unique immunoregulatory effects; 2) there is a definitive long-term (5 year) clinical benefit of the DBMC protocol on kidney allograft survival in which DBMC chimerism has expanded in the bone marrow compartment. In the recipient chimeric marrow, (recipient- derived) donor cells (RdD) and (recipient- derived) recipient cells (RdR) can be isolated and tested. There are sequentially increasing alloimmune inhibitory effects of RdD up to 2 years postoperatively, with phenotypic characterization of these long-term chimeric cells. DBMC has been cultured between 3 months and 1 year with recipient (allogeneic) feeder cells (DBMC-L), the latter an in vitro model somewhat analogous to RdD cell generation in vivo.
Our specific aims i n the present proposal are: I. A. To determine the influence of the modulating (immunoregulatory) DBMC, RdD, DBMC-L, and RdR cells on direct and indirect antigen presentation pathways. B. To analyze the capacity of those cells to induce specific T suppressor cells, i.e., infectious tolerance. C. To evaluate the effects of these cells on responding cell activation pathways, performing molecular analyses of TH1-TH2 cytokines. D. To analyze total cellular mRNA expression versus similarly treated spleen cells from the same cadaver donor by differential expression and cDNA array to delineate their unique immunoregulatory properties. E. To continue to follow up the recipients for levels of donor chimeric CD34+, CD3, CD19+ and other myeloid and lymphoid lineage cells by PCR Flow and cell isolation procedures, thereby correlating these studies with Specific Aim II. II. To continue to develop a reproducible multi-faceted protocol to monitor immunosuppressive withdrawal (now in progress) by protocol biopsy, limiting dilution analyses, alloantibody and cytokine ELISPOT studies, and trans-vivo assays in the rag-i knock-out mouse.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Project (R01)
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Surgery, Anesthesiology and Trauma Study Section (SAT)
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Flessner, Michael Francis
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University of Miami School of Medicine
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Leventhal, Joseph R; Miller, Joshua; Mathew, James M et al. (2018) Updated follow-up of a tolerance protocol in HLA-identical renal transplant pairs given donor hematopoietic stem cells. Hum Immunol 79:277-282
Leventhal, J R; Mathew, J M; Salomon, D R et al. (2016) Nonchimeric HLA-Identical Renal Transplant Tolerance: Regulatory Immunophenotypic/Genomic Biomarkers. Am J Transplant 16:221-34
Levitsky, Josh; Mathew, James M; Abecassis, Michael et al. (2013) Systemic immunoregulatory and proteogenomic effects of tacrolimus to sirolimus conversion in liver transplant recipients. Hepatology 57:239-48
Levitsky, Josh; Miller, Joshua; Huang, Xuemei et al. (2013) Inhibitory effects of belatacept on allospecific regulatory T-cell generation in humans. Transplantation 96:689-96
Leventhal, Joseph R; Mathew, James M; Salomon, Daniel R et al. (2013) Genomic biomarkers correlate with HLA-identical renal transplant tolerance. J Am Soc Nephrol 24:1376-85
Leventhal, J; Miller, J; Abecassis, M et al. (2013) Evolving approaches of hematopoietic stem cell-based therapies to induce tolerance to organ transplants: the long road to tolerance. Clin Pharmacol Ther 93:36-45
Leventhal, Joseph; Abecassis, Michael; Miller, Joshua et al. (2012) Chimerism and tolerance without GVHD or engraftment syndrome in HLA-mismatched combined kidney and hematopoietic stem cell transplantation. Sci Transl Med 4:124ra28
Levitsky, Josh; Leventhal, Joseph R; Miller, Joshua et al. (2012) Favorable effects of alemtuzumab on allospecific regulatory T-cell generation. Hum Immunol 73:141-9
Mathew, James M; Leventhal, Joseph R; Miller, Joshua (2011) Microchimerism in promoting graft acceptance in clinical transplantation. Curr Opin Organ Transplant 16:345-52
Yu, Yuming; Miller, Joshua; Leventhal, Joseph R et al. (2011) Requirement of cognate CD4+ T-cell recognition for the regulation of allospecific CTL by human CD4+ CD127- CD25+ FOXP3+ cells generated in MLR. PLoS One 6:e22450

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