HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints/Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Möbest, D. Articles by Henschler, R. Search for Related Content PubMed PubMed Citation Articles by Möbest, D. Articles by Henschler, R.

Differential Kinetics of Primitive Hematopoietic Cells Assayed In Vitro and In Vivo During Serum-Free Suspension Culture of CD34⁺ Blood Progenitor Cells

^a Experimental Hematology Group, Department of Hematology/Oncology, University Medical Center, Freiburg, Germany;
^b Department of Experimental Pharmacology, Max-Delbrück-Center for Molecular Medicine, Berlin, Germany;

Key Words. Hematopoietic progenitor cells • Serum-free medium • Transplantation • Ex vivo expansion

Dr. Reinhard Henschler, Department of Hematology/Oncology, University Medical Center, Hugstetter Strasse 55, 79106 Freiburg, Germany.

So far, blood progenitor cells (BPC) expanded ex vivo in the absence of stromal cells have not been demonstrated to reconstitute hematopoiesis in myeloablated patients. To characterize the fate of early hematopoietic progenitor cells during ex vivo expansion in suspension culture, human CD34⁺-enriched BPC were cultured in serum-free medium in the presence of FLT3 ligand (FL), stem cell factor (SCF) and interleukin 3 (IL-3). Both CD34 surface expression levels and the percentage of CD34⁺ cells were continuously downregulated during the culture period. We observed an expansion of colony-forming units granulocyte-macrophage (CFU-GM) and BFU-E beginning on day 3 of culture, reaching an approximate 2-log increase by days 5 to 7. Limiting dilution analysis of primitive in vitro clonogenic progenitors was performed through a week 6 cobblestone-area-forming cell (CAFC) assay, which has previously been shown to detect long-term bone marrow culture-initiating cells (LTC-IC). A maintenance or a slight (threefold) increase of week 6 CAFC/LTC-IC was found after one week of culture. To analyze the presence of BPC mediating in vivo engraftment, expanded CD34⁺ cells were transplanted into preirradiated NOD/SCID mice at various time points. Only CD34⁺ cells cultured for up to four days successfully engrafted murine bone marrow with human cells expressing myeloid or lymphoid progenitor phenotypes. In contrast, five- and seven-day expanded human BPC did not detectably engraft NOD/SCID mice. When FL, SCF and IL-3-supplemented cultures were performed for seven days on fibronectin-coated plastic, or when IL-3 was replaced by thrombopoietin, colony forming cells and LTC-IC reached levels similar to those of control cultures, yet no human cell engraftment was recorded in the mice. Also, culture in U-bottom microplates resulting in locally increased CD34⁺ cell density had no positive effect on engraftment. These results indicate that during ex vivo expansion of human CD34⁺ cells, CFC and LTC-IC numbers do not correlate with the potential to repopulate NOD/SCID mice. Our results suggest that ex vivo expanded BPC should be cultured for limited time periods only, in order to preserve bone-marrow-repopulating hematopoietic stem cells.

This article has been cited by other articles:

				H. M. Goselink, P. S. Hiemstra, P. van Noort, R. M.Y. Barge, R. Willemze, and J.H. F. Falkenburg Cytokine-Dependent Proliferation of Human CD34+ Progenitor Cells in the Absence of Serum Is Suppressed by Their Progeny's Production of Serine Proteinases Stem Cells, February 1, 2006; 24(2): 299 - 306. [Abstract] [Full Text] [PDF]

				O. Pelz, M. Wu, T. Nikolova, M. Kamprad, M. Ackermann, D. Egger, F. Emmrich, A. M. Wobus, and M. Cross Duplex Polymerase Chain Reaction Quantification of Human Cells in a Murine Background Stem Cells, June 1, 2005; 23(6): 828 - 833. [Abstract] [Full Text] [PDF]

				R. J. Ludwig, B. Boehme, M. Podda, R. Henschler, E. Jager, C. Tandi, W.-H. Boehncke, T. M. Zollner, R. Kaufmann, and J. Gille Endothelial P-Selectin as a Target of Heparin Action in Experimental Melanoma Lung Metastasis Cancer Res., April 15, 2004; 64(8): 2743 - 2750. [Abstract] [Full Text] [PDF]

				A. Nitsche, I. Junghahn, S. Thulke, J. Aumann, A. Radonic, I. Fichtner, and W. Siegert Interleukin-3 Promotes Proliferation and Differentiation of Human Hematopoietic Stem Cells but Reduces Their Repopulation Potential in NOD/SCID Mice Stem Cells, March 1, 2003; 21(2): 236 - 244. [Abstract] [Full Text] [PDF]

				J. P. Chute, A. A. Saini, D. J. Chute, M. R. Wells, W. B. Clark, D. M. Harlan, J. Park, M. K. Stull, C. Civin, and T. A. Davis Ex vivo culture with human brain endothelial cells increases the SCID-repopulating capacity of adult human bone marrow Blood, December 15, 2002; 100(13): 4433 - 4439. [Abstract] [Full Text] [PDF]

				T. Ro{beta}manith, B. Schroder, G. Bug, P. Muller, T. Klenner, R. Knaus, D. Hoelzer, and O.G. Ottmann Interleukin 3 Improves the Ex Vivo Expansion of Primitive Human Cord Blood Progenitor Cells and Maintains the Engraftment Potential of SCID Repopulating Cells Stem Cells, July 1, 2001; 19(4): 313 - 320. [Abstract] [Full Text] [PDF]