scholarly journals Differences among myeloproliferative disorders in the behavior of their restricted progenitor cells in culture

Blood ◽  
1983 ◽  
Vol 62 (3) ◽  
pp. 578-584 ◽  
Author(s):  
H Croizat ◽  
D Amato ◽  
DL McLeod ◽  
D Eskinazi ◽  
AA Axelrad
Keyword(s):  
Stem Cell ◽  
Progenitor Cells ◽  
Myeloproliferative Disorders ◽  
Hemopoietic Stem Cell ◽  
Erythroid Progenitor ◽  
Erythroid Progenitor Cells ◽  
Highly Sensitive ◽  
Common Observation ◽  
Normal Sensitivity ◽  
Hemopoietic Progenitor

Abstract We have studied the behavior in culture of circulating restricted hemopoietic progenitor cells from patients with idiopathic myelofibrosis (IMF), polycythemia vera (PV), and essential thrombocytopenia (ET). We have found differences in circulating granulocyte-macrophage, erythroid, and megakaryocytic progenitors that appear to be specific for these chronic myeloproliferative disorders. In IMF, most affected were granulocyte-macrophage progenitor cells (CFU- C), which circulated in increased numbers and were heterogeneous in their sensitivity to the regulatory factor(s) present in phytohemagglutinin (PHA) stimulated T-lymphocyte conditioned medium (CM). Most CFU-C were either highly sensitive to, or independent from, stimulatory factors, while others showed normal sensitivity. In some IMF patients, circulating megakaryocytic progenitors (CFU-M) were present that were capable of giving rise to colonies in the absence of added CM or erythropoietin (EPO). In PV, we confirmed the presence of circulating erythroid progenitor cells that give rise to colonies in culture without the addition of EPO. The number of circulating CFU-C was normal and they responded normally to CM. In ET, failure to detect 7-day circulating restricted progenitor cells was a common observation; the level of other circulating restricted progenitors was in the low normal range. Thus, despite certain common features, including a primary lesion at the level of the pluripotential hemopoietic stem cell, the myeloproliferative disorders differ with respect to the behavior in culture of their circulating restricted progenitor cells. These results have led us to postulate a second regulatory lesion in the pluripotential stem cell that differs in these disorders and is expressed at the level of the respective restricted progenitor cells.

scholarly journals Differences among myeloproliferative disorders in the behavior of their restricted progenitor cells in culture

Blood ◽  
1983 ◽  
Vol 62 (3) ◽  
pp. 578-584 ◽  
Author(s):  
H Croizat ◽  
D Amato ◽  
DL McLeod ◽  
D Eskinazi ◽  
AA Axelrad
Keyword(s):  
Stem Cell ◽  
Progenitor Cells ◽  
Myeloproliferative Disorders ◽  
Hemopoietic Stem Cell ◽  
Erythroid Progenitor ◽  
Erythroid Progenitor Cells ◽  
Highly Sensitive ◽  
Common Observation ◽  
Normal Sensitivity ◽  
Hemopoietic Progenitor

We have studied the behavior in culture of circulating restricted hemopoietic progenitor cells from patients with idiopathic myelofibrosis (IMF), polycythemia vera (PV), and essential thrombocytopenia (ET). We have found differences in circulating granulocyte-macrophage, erythroid, and megakaryocytic progenitors that appear to be specific for these chronic myeloproliferative disorders. In IMF, most affected were granulocyte-macrophage progenitor cells (CFU- C), which circulated in increased numbers and were heterogeneous in their sensitivity to the regulatory factor(s) present in phytohemagglutinin (PHA) stimulated T-lymphocyte conditioned medium (CM). Most CFU-C were either highly sensitive to, or independent from, stimulatory factors, while others showed normal sensitivity. In some IMF patients, circulating megakaryocytic progenitors (CFU-M) were present that were capable of giving rise to colonies in the absence of added CM or erythropoietin (EPO). In PV, we confirmed the presence of circulating erythroid progenitor cells that give rise to colonies in culture without the addition of EPO. The number of circulating CFU-C was normal and they responded normally to CM. In ET, failure to detect 7-day circulating restricted progenitor cells was a common observation; the level of other circulating restricted progenitors was in the low normal range. Thus, despite certain common features, including a primary lesion at the level of the pluripotential hemopoietic stem cell, the myeloproliferative disorders differ with respect to the behavior in culture of their circulating restricted progenitor cells. These results have led us to postulate a second regulatory lesion in the pluripotential stem cell that differs in these disorders and is expressed at the level of the respective restricted progenitor cells.

scholarly journals Stem cell factor retards differentiation of normal human erythroid progenitor cells while stimulating proliferation

Blood ◽  
1995 ◽  
Vol 86 (2) ◽  
pp. 572-580 ◽  
Author(s):  
K Muta ◽  
SB Krantz ◽  
MC Bondurant ◽  
CH Dai
Keyword(s):  
Stem Cell ◽  
Cell Viability ◽  
Progenitor Cells ◽  
Stem Cell Factor ◽  
Single Cells ◽  
Erythroid Differentiation ◽  
Tyrosine Kinase Receptor ◽  
Total Production ◽  
Erythroid Progenitor ◽  
Erythroid Progenitor Cells

Stem cell factor (SCF), the ligand for the c-kit tyrosine kinase receptor, markedly stimulates the accumulation of erythroid progenitor cells in vitro. We now report that SCF delays erythroid differentiation among the progeny of individual erythroid progenitors while greatly increasing the proliferation of these progeny. These effects appear to be independent of an effect on maintenance of cell viability. Highly purified day-6 erythroid colony-forming cells (ECFC), consisting mainly of colony-forming units-erythroid (CFU-E), were generated from human peripheral blood burst-forming units-erythroid (BFU-E). Addition of SCF to the ECFC in serum-free liquid culture, together with erythropoietin (EP) and insulin-like growth factor 1 (IGF-1), resulted in a marked increase in DNA synthesis, associated with a delayed peak in cellular benzidine positivity and a delayed incorporation of 59Fe into hemoglobin compared with cultures without SCF. In the presence of SCF, the number of ECFC was greatly expanded during this culture period, and total production of benzidine-positive cells plus hemoglobin synthesis were ultimately increased. To determine the effect of SCF on individual ECFC, single-cell cultures were performed in both semisolid and liquid media. These cultures demonstrated that SCF, in the presence of EP and IGF-1, acted on single cells and their descendants to delay erythroid differentiation while substantially stimulating cellular proliferation, without an enhancement of viability of the initial cells. This was also evident when the effect of SCF was determined using clones of ECFC derived from single BFU-E. Our experiments demonstrate that SCF acts on individual day-6 ECFC to retard erythroid differentiation while simultaneously providing enhanced proliferation by a process apparently independent of an effect on cell viability or programmed cell death.

scholarly journals Expression and function of receptors for stem cell factor and erythropoietin during lineage commitment of human hematopoietic progenitor cells

Blood ◽  
1996 ◽  
Vol 88 (5) ◽  
pp. 1594-1607 ◽  
Author(s):  
J Olweus ◽  
LW Terstappen ◽  
PA Thompson ◽  
F Lund-Johansen
Keyword(s):  
Stem Cell ◽  
Progenitor Cells ◽  
Stem Cell Factor ◽  
Progenitor Cell ◽  
High Sensitivity ◽  
Receptor Expression ◽  
Erythroid Progenitor ◽  
Erythroid Progenitor Cells ◽  
Fluorescence Measurements ◽  
And Function

The aim of the present study was to determine whether stem cell factor (SCF) and erythropoietin (EPO) act differently on defined subsets of progenitor cells, and if potential differences correlate with the receptor density on each subset. To investigate this possibility directly, we optimized conditions for the identification and purification of homogeneous progenitor cell subpopulations from human bone marrow. Populations containing 40% and 44% colony forming cells (CFCs) with 99% and 95% purity for the granulomonocytic and erythroid lineage, respectively, were sorted on the basis of differential expression of CD34, CD64, and CD71. In addition, a population containing 67% CFCs, of which 29–43% were CFU-MIX, was sorted from CD34hi CD38loCD50+ cells. Purified progenitor cell subsets were compared directly for responsiveness to SCF and EPO using a short-term proliferation assay. Expression of the receptors for SCF and EPO were then examined on each subset using a flow cytometer modified for high- sensitivity fluorescence measurements. The results show that EPO induces extensive proliferation of erythroid progenitor cells, but has no effect on the proliferation or survival of primitive or granulomonocytic progenitors, even when used in combination with other cytokines. The majority of erythroid progenitor cells furthermore stained positively with anti-EPO receptor (EPO-R) monoclonal antibodies, whereas other progenitor cells were negative. SCF alone induced extensive proliferation of erythroid progenitor cells, and had a stronger synergistic effect on primitive than on granulo-monocytic progenitors. In spite of these differences in SCF activity, there were no significant differences in SCF-R expression between the progenitor subsets. These results suggest that the selective action of EPO on erythropoiesis is determined by lineage-restricted receptor expression, whereas there are additional cell-type specific factors that influence progenitor cell responses to SCF.

A mouse model for visualization and conditional mutations in the erythroid lineage

Blood ◽  
2004 ◽  
Vol 104 (3) ◽  
pp. 659-666 ◽  
Author(s):  
Achim C. Heinrich ◽  
Roberta Pelanda ◽  
Ursula Klingmüller
Keyword(s):  
Stem Cell ◽  
Mouse Model ◽  
Progenitor Cells ◽  
Molecular Mechanisms ◽  
Regulatory Elements ◽  
Erythropoietin Receptor ◽  
Erythroid Progenitor ◽  
Hematopoietic Stem ◽  
Erythroid Progenitor Cells ◽  
Recombination System

AbstractHematologic disorders can be caused by sporadic or inherited mutations. However, the molecular mechanisms that lead to pathogenicity are only partially understood. An accurate method to generate mouse models is conditional gene manipulation facilitated by the Cre-loxP recombination system. To enable identification and genomic manipulation of erythroid progenitor cells, we established a knock-in mouse model (ErGFPcre) that expresses an improved GFPcre fusion protein controlled by the endogenous erythropoietin receptor (EpoR) promoter. We show that ErGFPcre mice enable the identification of GFP-positive erythroid progenitor cells and the highly specific genomic manipulation of the erythroid lineage. Analysis of GFP-positive erythroid progenitor cells suggests a developmental switch in lineage progression from the hematopoietic stem cell compartment to early erythroid progenitor cells that are stem cell antigen-1–negative (Sca-1–) and c-kithigh. Within the hematopoietic system, Cre-mediated recombination is limited to erythroid progenitor cells and occurs in the adult bone marrow at a frequency of up to 80% and in the fetal liver with an efficiency close to 100%. Differential transcriptional activity of the wild-type and the knock-in locus was observed in nonhematopoietic tissues. Thus, our ErGFPcre mouse model could promote the identification of regulatory elements controlling nonhematopoietic EpoR expression and facilitates the characterization and genomic manipulation of erythroid progenitor cells.

Expression and function of c-Kit in fetal hemopoietic progenitor cells: transition from the early c-Kit-independent to the late c-Kit-dependent wave of hemopoiesis in the murine embryo

Development ◽  
1993 ◽  
Vol 117 (3) ◽  
pp. 1089-1098 ◽  
Author(s):  
M. Ogawa ◽  
S. Nishikawa ◽  
K. Yoshinaga ◽  
S. Hayashi ◽  
T. Kunisada ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Progenitor Cells ◽  
Fetal Liver ◽  
Flow Cytometric Analysis ◽  
Yolk Sac ◽  
Erythroid Progenitor ◽  
Erythroid Progenitor Cells ◽  
And Function ◽  
Hemopoietic Progenitor ◽  
Hemopoietic Progenitor Cells

The protooncogene c-kit encodes a receptor type tyrosine kinase and is allelic with the W locus of mice. SLF, the c-Kit ligand which is encoded by the Sl locus, has growth promoting activity for hemopoietic stem cells. Previous studies demonstrated that c-Kit is functionally required for the proliferation of hemopoietic progenitor cells at various differentiation stages in adult bone marrow. However, the absence of functional SLF and c-Kit in fetuses with mutant alleles of Sl and W loci produces only minor effects on the myeloid and early erythroid progenitor cells in the fetal liver, although the level of the late erythroid progenitor cells is significantly affected. We used an anti-c-Kit monoclonal antibody to investigate the expression and function of c-Kit in murine fetal hemopoietic progenitor cells. Flow-cytometric analysis showed that hemopoiesis in the yolk sac and fetal liver started from cells that express c-Kit. The c-Kit expression decreased upon maturation into erythrocytes in each organ. By fluorescence activated cell sorting, the c-Kit+ cell population was enriched with the hemopoietic progenitor cells clonable in vitro (CFU-E, BFU-E and GM-CFC). To elucidate whether c-Kit functions in these progenitor cells in vivo, we took advantage of the antagonistic anti-c-Kit monoclonal antibody, ACK2, which can block the function of c-Kit. Administration of ACK2 after 12.5 days of gestation rapidly eliminated BFU-E and GM-CFC as well as CFU-E from the fetal liver. However, the number of these progenitor cells in the yolk sac and fetal liver was less affected when the fetuses were given ACK2 before 12.5 days of gestation. Our results provide evidence that there are two waves of hemopoiesis in murine embryos relative to c-Kit dependency. The c-Kit has an essential role on the growth of hemopoietic progenitor cells in the fetal liver after 12.5 days of gestation, whereas the progenitor cells in the liver and yolk sac of the earlier embryo do not depend on c-Kit and its ligand SLF.

scholarly journals Expression and function of receptors for stem cell factor and erythropoietin during lineage commitment of human hematopoietic progenitor cells

Blood ◽  
1996 ◽  
Vol 88 (5) ◽  
pp. 1594-1607 ◽  
Author(s):  
J Olweus ◽  
LW Terstappen ◽  
PA Thompson ◽  
F Lund-Johansen
Keyword(s):  
Stem Cell ◽  
Progenitor Cells ◽  
Stem Cell Factor ◽  
Progenitor Cell ◽  
High Sensitivity ◽  
Receptor Expression ◽  
Erythroid Progenitor ◽  
Erythroid Progenitor Cells ◽  
Fluorescence Measurements ◽  
And Function

Abstract The aim of the present study was to determine whether stem cell factor (SCF) and erythropoietin (EPO) act differently on defined subsets of progenitor cells, and if potential differences correlate with the receptor density on each subset. To investigate this possibility directly, we optimized conditions for the identification and purification of homogeneous progenitor cell subpopulations from human bone marrow. Populations containing 40% and 44% colony forming cells (CFCs) with 99% and 95% purity for the granulomonocytic and erythroid lineage, respectively, were sorted on the basis of differential expression of CD34, CD64, and CD71. In addition, a population containing 67% CFCs, of which 29–43% were CFU-MIX, was sorted from CD34hi CD38loCD50+ cells. Purified progenitor cell subsets were compared directly for responsiveness to SCF and EPO using a short-term proliferation assay. Expression of the receptors for SCF and EPO were then examined on each subset using a flow cytometer modified for high- sensitivity fluorescence measurements. The results show that EPO induces extensive proliferation of erythroid progenitor cells, but has no effect on the proliferation or survival of primitive or granulomonocytic progenitors, even when used in combination with other cytokines. The majority of erythroid progenitor cells furthermore stained positively with anti-EPO receptor (EPO-R) monoclonal antibodies, whereas other progenitor cells were negative. SCF alone induced extensive proliferation of erythroid progenitor cells, and had a stronger synergistic effect on primitive than on granulo-monocytic progenitors. In spite of these differences in SCF activity, there were no significant differences in SCF-R expression between the progenitor subsets. These results suggest that the selective action of EPO on erythropoiesis is determined by lineage-restricted receptor expression, whereas there are additional cell-type specific factors that influence progenitor cell responses to SCF.

scholarly journals Isolation of murine fetal hemopoietic progenitor cells and selective fractionation of various erythroid precursors

Blood ◽  
1981 ◽  
Vol 58 (2) ◽  
pp. 376-386 ◽  
Author(s):  
NA Nicola ◽  
D Metcalf ◽  
H von Melchner ◽  
AW Burgess
Keyword(s):  
Progenitor Cells ◽  
Fetal Liver ◽  
Pokeweed Mitogen ◽  
Erythroid Progenitor ◽  
Clonal Proliferation ◽  
Erythroid Progenitor Cells ◽  
Blast Cells ◽  
Hemopoietic Progenitor ◽  
Hemopoietic Progenitor Cells

Abstract Hemopoietic progenitor cells (colony- and cluster-forming cells in semisolid agar) were purified from light density CBA murine fetal liver cells using fluorescein-conjugated pokeweed mitogen (PWM) and a rhodamine-conjugated antineutrophil serum sandwich (alpha N) and three- parameter fluorescence-activated cell sorting. All clonable progenitor cells were highly enriched (36–50-fold) in PWM-positive (greater than channel 15), alpha N-negative (less than channel 30) fractions with relatively high intensity (greater than 100) low angle light scatter. No separation was achieved between different types of progenitor cells (granulocyte-macrophage and erythroid colony-forming cells). The enriched fraction was a pure population of large, basophilic, undifferentiated blast cells, and in agar cultures stimulated with colony-stimulating factors, up to 90% of the enriched cells were hemopoietic progenitor cells capable of varying levels of clonal proliferation. Further fractionation based on increasing fluorescence with PWM separated into discrete populations, nonproliferative morphologically recognizable erythroid cells, late erythroid progenitor cells (day 2 CFU-E), and cells forming pure or mixed erythroid burst colonies. In addition, the majority of pluripotential hemopoietic stem cells (CFU-SS) were clearly separated from progenitor cells forming colonies in vitro. The present techniques provide suitable numbers of enriched progenitor cells for a variety of biological and biochemical studies.

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