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Characterization and partial purification of human marrow cells capable of
initiating long-term hematopoiesis in vitro
HJ Sutherland, CJ Eaves, AC Eaves, W Dragowska and PM Lansdorp
Terry Fox Laboratory, B.C. Cancer Research Centre, Vancouver.
To develop a purification strategy for isolating the most primitive
hematopoietic stem cells present in normal human marrow we have combined
cell separation techniques with an assay for cells that initiate sustained
hematopoiesis in vitro in the presence of irradiated human marrow adherent
cells. These "feeders" were established by subculturing 2- to 6-week-old
primary long-term marrow culture adherent layers at a density of 3 x 10(4)
irradiated cells per square centimeter. Test "long-term culture
(LTC)-initiating cells" were plated on top of the feeders and the
cocultures then maintained as standard long-term marrow cultures with
half-media changes and removal of half of the nonadherent cells each week.
The total number of myeloid, erythroid, and multilineage clonogenic
progenitors present after 5 weeks was used to provide a quantitative
assessment of the number of LTC-initiating cells originally added. Using
this assay, the density, light scatter, and two cell surface antigen
properties of LTC- initiating cells have been defined and compared with
cells capable of directly forming colonies in methylcellulose. While the
majority of the clonogenic cells were found in the high forward light
scatter (FLS) "blast" window, LTC-initiating cells had significantly lower
FLS properties and in this respect were more similar to lymphocytes. LTC-
initiating cells also expressed less HLA-DR antigen than clonogenic cells.
The majority of LTC-initiating cells were found in the top 2% of the CD34
(My10) fluorescence profile, whereas clonogenic cells were found throughout
the top 5% of the CD34 fluorescence profile. By combining low FLS, low
orthogonal light scatter (OLS), low HLA-DR expression, and high CD34
expression, a population could be obtained that was enriched for
LTC-initiating cells approximately 800-fold over unseparated marrow. This
population contains only 0.06% of the marrow cells and 2% of the total
clonogenic cells, but retains 50% to 60% of the LTC-initiating cells
present in the original marrow. The ability to purify these two populations
independently shows that the LTC and clonogenic assays identify distinct,
although not necessarily nonoverlapping cell types in human marrow. Since
clonogenic cells are derived from LTC-initiating cells, the LTC assay
clearly detects a more primitive population. The availability of a simple
approach that allows the purification of such cells by three orders of
magnitude in high yield should be useful for the investigation of early
events in hematopoiesis as well as for the definitive isolation of human
hematopoietic stem cells with long-term in vivo repopulating potential.
Volume 74,
Issue 5,
pp. 1563-1570,
10/01/1989
Copyright © 1989 by The American Society of Hematology
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J. C.Y. Wang, M. Doedens, and J. E. Dick
Primitive Human Hematopoietic Cells Are Enriched in Cord Blood Compared With Adult Bone Marrow or Mobilized Peripheral Blood as Measured by the Quantitative In Vivo SCID-Repopulating Cell Assay
Blood,
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[Abstract]
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S. K. Nilsson, M. S. Dooner, C. Y. Tiarks, H.-U. Weier, and P. J. Quesenberry
Potential and Distribution of Transplanted Hematopoietic Stem Cells in a Nonablated Mouse Model
Blood,
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[Abstract]
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A. C. Berardi, E. Meffre, F. Pflumio, A. Katz, W. Vainchenker, C. Schiff, and L. Coulombel
Individual CD34+CD38lowCD19-CD10- Progenitor Cells From Human Cord Blood Generate B Lymphocytes and Granulocytes
Blood,
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3554 - 3564.
[Abstract]
[Full Text]
[PDF]
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A. Blair, D.E. Hogge, L.E. Ailles, P.M. Lansdorp, and H.J. Sutherland
Lack of Expression of Thy-1 (CD90) on Acute Myeloid Leukemia Cells With Long-Term Proliferative Ability In Vitro and In Vivo
Blood,
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89(9):
3104 - 3112.
[Abstract]
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C. Sanz, A. Benito, M. Silva, B. Albella, C. Richard, J. C. Segovia, A. Insunza, J. A. Bueren, and J. L. Fernandez-Luna
The Expression of Bcl-x Is Downregulated During Differentiation of Human Hematopoietic Progenitor Cells Along the Granulocyte But Not the Monocyte/Macrophage Lineage
Blood,
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89(9):
3199 - 3204.
[Abstract]
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A. J. Szczepek, P.L. Bergsagel, L. Axelsson, C. B. Brown, A. R. Belch, and L. M. Pilarski
CD34+ Cells in the Blood of Patients With Multiple Myeloma Express CD19 and IgH mRNA and Have Patient-Specific IgH VDJ Gene Rearrangements
Blood,
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[Abstract]
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J. P. Maciejewski, E. M. Sloand, T. Sato, S. Anderson, and N. S. Young
Impaired Hematopoiesis in Paroxysmal Nocturnal Hemoglobinuria/Aplastic Anemia Is Not Associated With a Selective Proliferative Defect in the Glycosylphosphatidylinositol-Anchored Protein-Deficient Clone
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[Abstract]
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R. M. Lemoli, A. Tafuri, A. Fortuna, M. T. Petrucci, M. R. Ricciardi, L. Catani, D. Rondelli, M. Fogli, G. Leopardi, C. Ariola, et al.
Cycling Status of CD34+ Cells Mobilized Into Peripheral Blood of Healthy Donors by Recombinant Human Granulocyte Colony-Stimulating Factor
Blood,
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[Abstract]
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K. Inoue, H. Ogawa, Y. Sonoda, T. Kimura, H. Sakabe, Y. Oka, S. Miyake, H. Tamaki, Y. Oji, T. Yamagami, et al.
Aberrant Overexpression of the Wilms Tumor Gene (WT1) in Human Leukemia
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[Abstract]
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A. Aiuti, I.J. Webb, C. Bleul, T. Springer, and J.C. Gutierrez-Ramos
The Chemokine SDF-1 Is a Chemoattractant for Human CD34+ Hematopoietic Progenitor Cells and Provides a New Mechanism to Explain the Mobilization of CD34+ Progenitors to Peripheral Blood
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R. Mohle, M. A.S. Moore, R. L. Nachman, and S. Rafii
Transendothelial Migration of CD34+ and Mature Hematopoietic Cells: An In Vitro Study Using a Human Bone Marrow Endothelial Cell Line
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J.-P. Laporte, N.-C. Gorin, P. Rubinstein, S. Lesage, M.-F. Portnoi, V. Barbu, M. Lopez, L. Douay, and A. Najman
Cord-Blood Transplantation from an Unrelated Donor in an Adult with Chronic Myelogenous Leukemia
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