|
|
 Previous Article | Table of Contents | Next Article 
Cycle initiation and colony formation in culture by murine marrow cells
with long-term reconstituting potential in vivo
M Trevisan, XQ Yan and NN Iscove
Ontario Cancer Institute, Toronto, Canada.
This investigation was directed at separating long-term reconstituting
(LTR) stem cells in normal murine marrow from hematopoietic precursors
detectable in short-term assays in vitro and in vivo, and then at
determining whether purified LTR cells could themselves form colonies in
culture. To do so, it was first necessary to identify culture conditions
that would induce their growth while preserving their long- term
reconstituting capacity. Marrow was cultured with various cytokines in
liquid suspension for 4 days, after which the surviving LTR activity was
quantitated in a competitive in vivo assay. Activity was preserved near
input levels with combined murine c-kit ligand (KL), interleukin-1 (IL-1),
IL-6, and IL-11. When the cultures also included tritiated or unlabeled
thymidine, LTR potential was eliminated, indicating that essentially all
LTR cells were induced into cell cycle with these cytokines. To purify
them, marrow was sorted on the basis of Ly6A expression and Rhodamine 123
retention. The Ly6AhiRh123ls fraction contained 85% of total recovered LTR
activity but only 1% of the recovered cells measured by multilineage colony
formation in spleens or in vitro. This fraction was cultured in methyl
cellulose with KL, IL-1, IL-6, and IL-11 for 4 to 6 days, after which
colonies were isolated and injected into mice. High levels of permanent
reconstitution were achievable in sublethally irradiated W41/W41 mice after
the injection of a single reconstituting unit, and limiting dilution
analysis estimated the frequency of multilineage LTR at 1 in 11,200
unpurified adult marrow cells. In either lethally irradiated normal or
sublethally irradiated W41/W41 mice, 1-year lymphomyeloid reconstitutions
were obtained from 1 in 65 to 84 colonies of 2 to 16 dispersed cells, but
not from larger colonies or those with clumped cells. The results establish
that resting marrow LTR cells can be separated from almost all of the more
advanced clonogenic cells that are still pluripotential, can be induced to
cycle in culture by defined cytokines with preservation of their
reconstituting potential, and can be manipulated and assayed efficiently at
single-cell and colony levels.
Volume 88,
Issue 11,
pp. 4149-4158,
12/01/1996
Copyright © 1996 by The American Society of Hematology
 CiteULike  Connotea  Del.icio.us  Digg  Reddit  Technorati What's this?
This article has been cited by other articles:
|
|
|
|
 
R. H. Cho, H. B. Sieburg, and C. E. Muller-Sieburg
A new mechanism for the aging of hematopoietic stem cells: aging changes the clonal composition of the stem cell compartment but not individual stem cells
Blood,
June 15, 2008;
111(12):
5553 - 5561.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
D. Kent, M. Copley, C. Benz, B. Dykstra, M. Bowie, and C. Eaves
Regulation of Hematopoietic Stem Cells by the Steel Factor/KIT Signaling Pathway
Clin. Cancer Res.,
April 1, 2008;
14(7):
1926 - 1930.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
H. B. Sieburg, R. H. Cho, B. Dykstra, N. Uchida, C. J. Eaves, and C. E. Muller-Sieburg
The hematopoietic stem compartment consists of a limited number of discrete stem cell subsets
Blood,
March 15, 2006;
107(6):
2311 - 2316.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
N. Uchida, B. Dykstra, K. Lyons, F. Leung, M. Kristiansen, and C. Eaves
ABC transporter activities of murine hematopoietic stem cells vary according to their developmental and activation status
Blood,
June 15, 2004;
103(12):
4487 - 4495.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. E. Muller-Sieburg, R. H. Cho, M. Thoman, B. Adkins, and H. B. Sieburg
Deterministic regulation of hematopoietic stem cell self-renewal and differentiation
Blood,
July 30, 2002;
100(4):
1302 - 1309.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
F. Billia, M. Barbara, J. McEwen, M. Trevisan, and N. N. Iscove
Resolution of pluripotential intermediates in murine hematopoietic differentiation by global complementary DNA amplification from single cells: confirmation of assignments by expression profiling of cytokine receptor transcripts
Blood,
April 15, 2001;
97(8):
2257 - 2268.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. Domen and I. L. Weissman
Hematopoietic Stem Cells Need Two Signals to Prevent Apoptosis; Bcl-2 Can Provide One of These, Kitl/C-KIT Signaling the Other
J. Exp. Med.,
December 18, 2000;
192(12):
1707 - 1718.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
V. Maguer-Satta, R. Oostendorp, D. Reid, and C. J. Eaves
Evidence that ceramide mediates the ability of tumor necrosis factor to modulate primitive human hematopoietic cell fates
Blood,
December 15, 2000;
96(13):
4118 - 4123.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
S. Hatada, K. Nikkuni, S. A. Bentley, S. Kirby, and O. Smithies
Gene correction in hematopoietic progenitor cells by homologous recombination
PNAS,
November 16, 2000;
(2000)
240462897.
[Abstract]
[Full Text]
|
|
|
|
|
|
|
D. Bryder and S. E. W. Jacobsen
Interleukin-3 supports expansion of long-term multilineage repopulating activity after multiple stem cell divisions in vitro
Blood,
September 1, 2000;
96(5):
1748 - 1755.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
P. W. Zandstra, D. A. Lauffenburger, and C. J. Eaves
A ligand-receptor signaling threshold model of stem cell differentiation control: a biologically conserved mechanism applicable to hematopoiesis
Blood,
August 15, 2000;
96(4):
1215 - 1222.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
R. A. J. Oostendorp, J. Audet, and C. J. Eaves
High-resolution tracking of cell division suggests similar cell cycle kinetics of hematopoietic stem cells stimulated in vitro and in vivo
Blood,
February 1, 2000;
95(3):
855 - 862.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
G. Guenechea, J.C. Segovia, B. Albella, M. Lamana, M. Ramirez, C. Regidor, M.N. Fernandez, and J.A. Bueren
Delayed Engraftment of Nonobese Diabetic/Severe Combined Immunodeficient Mice Transplanted With Ex Vivo-Expanded Human CD34+ Cord Blood Cells
Blood,
February 1, 1999;
93(3):
1097 - 1105.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
L. Maulon, S. Guérin, J. E. Ricci, Y. Colin, J. P. Cartron, and P. Auberger
Differential expression of the Kell blood group and CD10 antigens: two related membrane metallopeptidases during differentiation of K562 cells by phorbol ester and hemin
FASEB J,
May 1, 1998;
12(7):
531 - 539.
[Abstract]
[Full Text]
|
|
|
|
|
|
|
C. L. Miller and C. J. Eaves
Expansion in vitro of adult murine hematopoietic stem cells with transplantable lympho-myeloid reconstituting ability
PNAS,
December 9, 1997;
94(25):
13648 - 13653.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
E. Conneally, J. Cashman, A. Petzer, and C. Eaves
Expansion in vitro of transplantable human cord blood stem cells demonstrated using a quantitative assay of their lympho-myeloid repopulating activity in nonobese diabetic-scid/scid mice
PNAS,
September 2, 1997;
94(18):
9836 - 9841.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
S. Hatada, K. Nikkuni, S. A. Bentley, S. Kirby, and O. Smithies
Gene correction in hematopoietic progenitor cells by homologous recombination
PNAS,
December 5, 2000;
97(25):
13807 - 13811.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
