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Migratory Pathways and Engraftment of Human Cord Blood Derived Ex-vivo Created Di- Chimeric Cells and its Association with Therapeutic Effect on the Allograft Survival.
Joanna Cwykiel, MS1, Medhat Askar, MD PHD2, Grzegorz Kwiecien, MD2, Maria Siemionow, MD PHD DSc1.
1University of Illinois at Chicago, Chicago, IL, USA, 2Cleveland Clinic, Cleveland, OH, USA.

PURPOSE:
To evaluate the survival, migratory pathways and engraftment of ex-vivo fused human cord blood derived di-chimeric cells (DCC) in the nude rat model.
METHODS:
Twenty-four fusions of human umbilical cord blood (UCB) cells were performed. UCB mononuclear cells from 2 donors were stained separately with PKH26 and PKH67 dyes. Fusion was performed using polyethylene glycol (PEG). Double (PKH26/ PKH67) stained DCC were sorted and subjected to lymphocytotoxicity (LCT) test and STR-PCR. DCC (4-9x106) were injected to the femur of rat recipients. The presence of DCC was assessed in 6 groups at: Group1 - 24hours, Group2 - 72hours, Group3 - 7days, Group4 - 21days, Group5 - 1month and Group6 - 2months. The presence of DCC was assessed in blood, bone marrow (BM), lymph nodes, spleen, lung, liver, skin and brain using flow cytometry (FC) and immunofluorescent staining.
RESULTS:
DCC creation was confirmed using FC and microscopy. LCT analysis showed HLA class I and II from both UCB donors present on the surface of DCC. The engraftment of DCC was detected by anti-human HLA class I staining. Study indicated that DCC within 24 hours were capable of migrating from the injected to contralateral bone as well as to the lymphoid organs such as spleen and lymph nodes and were observed up to 2-months after delivery.
CONCLUSION:
The presence of DCC in BM compartment and lymphoid tissues of the recipient confirmed DCC engraftment and long-term survival. The unique concept of DCC therapy introduces novel applications in solid organ and VCA transplantation.


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