Faculty Collaboration Database

Ex vivo expansion of megakaryocyte precursors from umbilical cord blood CD34 cells in a closed liquid culture system. Biol Blood Marrow Transplant 2003 Mar;9(3):151-6

Date

03/26/2003

Pubmed ID

12652464

DOI

10.1053/bbmt.2003.50013

Scopus ID

2-s2.0-0642339113 (requires institutional sign-in at Scopus site)   20 Citations

Abstract

Umbilical cord blood (UCB) provides a rich source of stem cells for transplantation after myeloablative therapy. One major disadvantage of UCB transplantation is delayed platelet engraftment. We propose to hasten platelet engraftment by expanding the number of megakaryocyte (MK) precursors (CD34/CD41 cells) through cytokine stimulation within a closed, pre-clinical liquid culture system. Clinical engraftment data suggest a 5- to 10-fold increase in MK precursors in a UCB unit can accelerate platelet engraftment, so this was our goal. Thirteen UCB samples from full-term births were Ficoll-separated and frozen for subsequent use. On thawing, the mononuclear cell population was positively selected for CD34(+) expression. The cells were cultured in gas-permeable Teflon-coated bags in serum-free medium containing the following cytokines: recombinant human interleukin-3, recombinant human Flt3 ligand, recombinant human stem cell factor, and recombinant human thrombopoietin. MK lineage cell expansion was assessed using mononuclear cell count and flow cytometry (CD34/41, CD41, CD34/61, and CD61 expression) on days 7, 11, and 14. Optimal expansion of CD34/41 and CD41 cells was observed at day 11, with a median 6-fold and 33-fold increase in the starting cell doses, respectively. CD34/61 and CD61 cell expansion at day 11 was 7-fold and 14-fold, respectively. MK precursors can be successfully expanded from CD34(+) UCB cells in a closed liquid culture system using interleukin-3, recombinant human Flt3 ligand, recombinant human stem cell factor, and recombinant human thrombopoietin to a level that should have a clinical impact in the transplantation setting. Our ex vivo expansion technique needs to be further optimized before it can be used in a pilot UCB transplantation trial.

Author List

Shaw PH, Gilligan D, Wang XM, Thall PF, Corey SJ

Author

Peter H. Shaw MD Professor in the Pediatrics department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

Antigens, CD
Antigens, CD34
Cell Culture Techniques
Cell Division
Culture Media, Serum-Free
Cytokines
Erythroid Precursor Cells
Fetal Blood
Humans
Megakaryocytes
Models, Statistical