Date: Sunday, May 3, 2015
Session Time: 5:30pm-6:30pm
Presentation Time: 5:30pm-6:30pm
Location: Exhibit Hall E
PURPOSE: The aim of this study was to evaluate safety, survival, migration and engraftment of ex-vivo fused human cord blood derived di-chimeric cells (DCC) in the NOD SCID mouse model.
METHODS: A total of 22 fusions of human umbilical cord blood (UCB) cells were performed. UCB from 2 unrelated donors were stained with PKH26 and PKH67 dyes. Cell fusion, performed with polyethylene glycol was confirmed by double (PKH26/ PKH67) staining of DCC which were sorted and subjected to the following in vitro evaluations (10 fusions): lymphocytotoxicity (LCT) test, PCR-SSOP, STR-PCR, viability (Annexin-V, Tunel, LIVE/DEAD staining), phenotype, colony forming unit (CFU) and COMET assay. DCC (3-5×106 cells) from 12 fusions were delivered: Group 1: intraosseous (n=4), Group 2: intramuscular (n=4) or Group 3: subcutaneous (n=4) to the recipient mice. Control mice (n=12) received 2-3×106 of UCB delivered intraosseous, intramuscular or subcutaneous. Mice were evaluated by palpation (daily) and histology (end-point) for the tumor growth. Tumor formation in Group 1 was tested bi-monthly by X-ray. DCC migratory pathways were assessed in blood, bone marrow, lymph nodes, spleen, lung, and liver at 4 months after delivery using anti-human HLA class I staining and PCR.
RESULTS: LCT showed HLA class I and II from both UCB donors on the surface of DCC and results were confirmed by PCR-SSOP and PCR-STR. Annexin V, Tunel and Live/Dead staining revealed limited effect of fusion on the viability of DCC. CFU determined proliferative properties of DCC comparable to UCB. COMET assay confirmed no damage to the DNA of DCC. Clinical observation and X-ray did not detect tumor formation in any group. In Group 3, migratory properties of DCC were confirmed at 24 and 72 hours after cell delivery by detection of DCC in the blood. Immunofluorescent, histological and PCR analysis are currently performed.
CONCLUSIONS: We successfully characterized phenotype and confirmed viability, proliferation, safety and migratory properties of DCC. The unique concept of DCC supportive therapy, introducing cells presenting phenotype characteristic of both transplant donor and recipient is a new approach for development of transplant tolerance induction strategy.
To cite this abstract in AMA style:Cwykiel J, Sklodowska A, Malewski W, Cyran M, Karagoz H, Tfaily EBryndza, Askar M, Siemionow M. Phenotype Characterization, Migratory Pathways, Engraftment and Safety Evaluation of Human Cord Blood Derived Ex-Vivo Created Di- Chimeric Cells in NOD SCID Mouse Model: A Preliminary Study [abstract]. Am J Transplant. 2015; 15 (suppl 3). https://atcmeetingabstracts.com/abstract/phenotype-characterization-migratory-pathways-engraftment-and-safety-evaluation-of-human-cord-blood-derived-ex-vivo-created-di-chimeric-cells-in-nod-scid-mouse-model-a-preliminary-study/. Accessed August 4, 2020.
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