Role of cytokines in sustaining long-term human megakaryocytopoiesis in vitro.
An in vitro liquid suspension culture system was used to determine the role of cytokines in sustaining long-term human megakaryocytopoiesis. Bone marrow cells expressing CD34 but not HLA-DR (CD34+DR-) were used as the inoculum of cells to initiate long-term bone marrow cultures (LTBMC). CD34+DR- cells (5 x 10(3)/mL) initially contained 0.0 +/- 0.0 assayable colony-forming unit-megakaryocytes (CFU-MK), 6.2 +/- 0.4 assayable burst-forming unit-megakaryocytes (BFU-MK), and 0.0 +/- 0.0 megakaryocytes (MK). LTBMCs were recharged every 48 hours with granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-1 alpha (IL-1 alpha), IL-3, and/or IL-6, alone or in combination. LTBMCs were demidepopulated weekly or biweekly, the number of cells and MK enumerated, and then assayed for CFU-MK and BFU-MK. LTBMCs receiving no cytokine(s) contained no assayable CFU-MK or BFU-MK and no observable MK. LTBMCs receiving GM-CSF, IL-1 alpha, and/or IL-3 contained assayable CFU-MK and MK but no BFU-MK for 10 weeks of culture. The effects of GM-CSF and IL-3, IL-1 alpha and IL-3, but not GM-CSF and IL-1 alpha were additive with regards to their ability to augment the numbers of assayable CFU-MK during LTBMC. LTBMCs supplemented with IL-6 contained modest numbers of assayable CFU-MK for only 4 weeks; this effect was not additive to that of GM-CSF, IL-1 alpha, or IL-3. The addition of GM-CSF, IL-1 alpha, and IL-3 alone or in combination each led to the appearance of significant numbers of MKs during LTBMC. By contrast, IL-6 supplemented cultures contained relatively few MK. These studies suggest that CD34+DR- cells are capable of initiating long-term megakaryocytopoiesis in vitro and that a hierarchy of cytokines exists capable of sustaining this process.
Department of Medicine, Indiana University School of Medicine, Indianapolis.
R A
RA
Briddell
J E
JE
Brandt
T B
TB
Leemhuis
R
R
Hoffman
D015703:Antigens, CD / Q000032:analysis; D018952:Antigens, CD34; D001854:Bone Marrow Cells; D002478:Cells, Cultured; D003114:Colony-Forming Units Assay; D016207:Cytokines / Q000494:pharmacology*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006684:HLA-DR Antigens / Q000032:analysis; D006410:Hematopoiesis; D006412:Hematopoietic Stem Cells / Q000166:cytology; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology; D007377:Interleukin-3 / Q000494:pharmacology; D015850:Interleukin-6 / Q000494:pharmacology; D008533:Megakaryocytes / Q000166:cytology* / Q000276:immunology
D015703:Antigens, CD; D018952:Antigens, CD34; D016207:Cytokines; D006684:HLA-DR Antigens; D007375:Interleukin-1; D007377:Interleukin-3; D015850:Interleukin-6; D016178:Granulocyte-Macrophage Colony-Stimulating Factor