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Cytomatrix |
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Expansion of Cord Blood Progenitors in a Novel 3-Dimensional Bioreactor | ||
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Bettina M Ehring, Daniel I Plosky, Kristen L Biber, Julie N Reitter,
Cytomatrix, Woburn, MA; Karen Ballen, University of Mass., Worcester, MA; Mark Poposky, American Red Cross, Dedham, MA; Mark J Pykett, Michael Rosenzweig, Cytomatrix, Woburn, MA |
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Background: Umbilical cord blood (UCB) has been demonstrated as a valuable source of hematopoietic progenitors for allogeneic stem cell transplantation. Due to the size of the potential graft these cells have only provided an effective source of stem cells for individuals of <40kg. An obvious limitation in achieving universal availability of cord progenitors is a means to expand the number of cells required for hematopoietic reconstitution in an adult. To date, efforts to expand UCB progenitors have used large quantities of cytokines and although the number of cells expanded in vitro, graft recipients have demonstrated variable responses post-transplant that have included profoundly delayed platelet engraftment and a higher incidence of GVHD. We have examined alternative strategies of UCB expansion that promote proliferation of primitive progenitors with limited differentiation. We have previously described the utility of a three-dimensional matrix, CellfoamTM, for the serum-free, cytokine-free culture and expansion of both bone-marrow and peripheral blood mobilized stem cells (Bagley et al, Exp. Hem.1998). In this study we investigated the performance of this culture device to promote expansion of UCB progenitors in the absence of serum and cytokines. Methods: Fresh cord blood samples (n=9) were centrifuged over a Ficoll gradient prior to CD34 separation with immunomagnetic beads. CD34+ cells with a purity of 27- 73% were cultured in the CellfoamTM device for 2 weeks in serum-free media without the addition of exogenous cytokines. Cells were harvested after 2 weeks and examined by flow cytometry. Results: Evaluation of CD34 expression in multiple replicates demonstrated a 2-4 fold increase in the number of CD45+CD34+ cells during the culture period. The number of CD34+CD38- cells increased by 2-3 logs above baseline during this time frame. The increase in both CD34+ and CD34+38- cells above baseline is significant (p<0.001). To determine the function of cells recovered from Cellfoam cultures we used a standard NOD/SCID transplant model. Data demonstrated that these cultured, expanded cells retained the capacity to engraft in vivo. Conclusion: This data demonstrates the efficacy of CellfoamTM as a three-dimensional tissue matrix that may enable a means to effectively expand UCB progenitors in a way that maintains pluripotency of the graft as well as achieving a cell dose that is clinically useful for adults.   |
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