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Developing a co-culture system for effective megakaryo/thrombopoiesis from umbilical cord blood hematopoietic stem/progenitor cells.

AbstractBACKGROUND AIMS:
Platelet transfusion can be a life-saving procedure in different medical settings. Thus, there is an increasing demand for platelets, of which shelf-life is only 5 days. The efficient ex vivo biomanufacturing of platelets would allow overcoming the shortages of donated platelets.
METHODS:
We exploited a two-stage culture protocol aiming to study the effect of different parameters on the megakaryo/thrombopoiesis ex vivo. In the expansion stage, human umbilical cord blood (UCB)-derived CD34(+)-enriched cells were expanded in co-culture with human bone marrow mesenchymal stromal cells (BM-MSCs). The megakaryocytic commitment and platelet generation were studied, considering the impact of exogenous addition of thrombopoietin (TPO) in the expansion stage and a cytokine cocktail (Cyt) including TPO and interleukin-3 in the differentiation stage, with the use of different culture medium formulations, and in the presence/absence of BM-MSCs (direct versus non-direct cell-cell contact).
RESULTS:
Our results suggest that an early megakaryocytic commitment, driven by TPO addition during the expansion stage, further enhanced megakaryopoiesis. Importantly, the results suggest that co-culture with BM-MSCs under serum-free conditions combined with Cyt addition, in the differentiation stage, significantly improved the efficiency yield of megakaryo/thrombopoiesis as well as increasing %CD41, %CD42b and polyploid content; in particular, direct contact of expanded cells with BM-MSCs, in the differentiation stage, enhanced the efficiency yield of megakaryo/thrombopoiesis, despite inhibiting their maturation.
CONCLUSIONS:
The present study established an in vitro model for the hematopoietic niche that combines different biological factors, namely, the presence of stromal/accessory cells and biochemical cues, which mimics the BM niche and enhances an efficient megakaryo/thrombopoiesis process ex vivo.
AuthorsJavad Hatami, Pedro Z Andrade, António Pedro Alves de Matos, Dusan Djokovic, Carla Lilaia, Frederico Castelo Ferreira, Joaquim M S Cabral, Cláudia L da Silva
JournalCytotherapy (Cytotherapy) Vol. 17 Issue 4 Pg. 428-42 (Apr 2015) ISSN: 1477-2566 [Electronic] England
PMID25680300 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
CopyrightCopyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.
Chemical References
  • Antigens, CD34
  • IL3 protein, human
  • Interleukin-3
  • Thrombopoietin
Topics
  • Antigens, CD34 (metabolism)
  • Blood Platelets (cytology)
  • Cell Communication (physiology)
  • Cell Differentiation (drug effects)
  • Cell Proliferation (drug effects)
  • Cells, Cultured
  • Coculture Techniques
  • Fetal Blood (cytology)
  • Hematopoietic Stem Cells (cytology)
  • Humans
  • Interleukin-3 (pharmacology)
  • Megakaryocytes (cytology)
  • Mesenchymal Stem Cells (cytology)
  • Platelet Transfusion (methods)
  • Thrombopoiesis (physiology)
  • Thrombopoietin (pharmacology)

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