Human megakaryocytic progenitors (CFU-M) assayed in methylcellulose: Physical characteristics and requirements for growth

H. Kimura, S. A. Burstein, D. Thorning, Jerry S Powell, L. A. Harker, P. J. Fialkow, J. W. Adamson

Research output: Contribution to journalArticlepeer-review

54 Scopus citations

Abstract

The basic culture requirements and several physical characteristics were defined for megakaryocytic colony-forming cells (CFU-M) from normal human marrow growing in methylcellulose. Ficoll-hypaque separated mononuclear cells from human marrow gave rise to megakaryocytic colonies in the presence of human plasma and phytohemagglutinin-stimulated leukocyte-conditioned medium (PHA-LCM). Their identity as megakaryocytic colonies was confirmed by immunofluorescence staining with a monoclonal antibody to human factor VIII antigen and by electron microscopy of individually harvested colonies. Demonstration of the single-cell origin of the colonies was provided by analysis of the glucose-6-phosphate dehydrogenase (G-6-PD) enzyme type of individually harvested colonies grown from a G-6-PD heterozygote. The colonies grew best in heparinized or citrated plasma as opposed to serum. Detailed studies suggested that platelet-release products were responsible for this difference. Tritiated thymidine suicide studies showed that the percentage of CFU-M in DNA synthesis was 23 ± 8% (n = 10). The modal velocity sedimentation rate of CFU-M was 4.9 ± 0.6 mm/hr (n = 4) while that of concurrently studied granulocyte/macrophage colony-forming cells (CFU-GM) was 5.7 ± 0.5 mm/hr. Examination of the PHA-LCM dose-response characteristics suggested the presence in the conditioned medium of an inhibitor to megakaryocyte colony growth which was partially removed by chromatography of the medium on Sephadex G-100. The resulting conditioned medium increased the cloning efficiency for CFU-M compared with that with crude PHA-LCM (15.3 ± 7.0 and 8.2 ± 5.3/105 marrow cells, respectively).

Original languageEnglish (US)
Pages (from-to)87-96
Number of pages10
JournalJournal of Cellular Physiology
Volume118
Issue number1
StatePublished - 1984
Externally publishedYes

ASJC Scopus subject areas

  • Clinical Biochemistry
  • Cell Biology
  • Physiology

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