scholarly journals Mammalian Homeobox B6 Expression Can Be Correlated With Erythropoietin Production Sites and Erythropoiesis During Development, But Not With Hematopoietic or Nonhematopoietic Stem Cell Populations

Blood ◽  
1997 ◽  
Vol 89 (8) ◽  
pp. 2723-2735 ◽  
Author(s):  
Frank Zimmermann ◽  
Ivan N. Rich
Keyword(s):  
Gene Expression ◽  
Stem Cell ◽  
Hox Genes ◽  
Fetal Liver ◽  
Single Gene ◽  
Lymphoid Cells ◽  
Cell Populations ◽  
Hematopoietic Stem ◽  
Hox Gene ◽  
Stem Cell Populations

Abstract There has been increasing interest in the involvement of mammalian homeobox (HOX) genes in hematopoietic regulation. The HOX genes are clustered in 4 chromosomes in mice and humans. In general, 5′ end HOX gene expression is predominant in hematopoietic stem cell populations, whereas 3′ end HOX gene expression are primarily found in committed progenitor cells. Furthermore, HOX genes of the A cluster are generally found in myelomonocytic cells, B cluster genes in erythropoietic cells, and C cluster genes in lymphoid cells. The results presented here concentrate on a single gene, namely HOX B6. Preliminary observations using whole mount in situ hybridization showed that both HOX B6 and erythropoietin (EPO) gene expression occurred in exactly the same areas of the 8.5-day mouse embryo. As a consequence, we studied the expression of HOX B6 and EPO gene expression from 6.5 to 19.5 days of gestation, in the neonate, and in the adult. It was found that the sequential transfer of erythropoiesis in different organs during development was followed by a similar transfer of HOX B6 and EPO gene expression. Between days 16.5 and 17.5, both HOX B6 and EPO gene expression decrease in the fetal liver, even though hepatic erythropoiesis continues to decline and is transferred to the fetal spleen. Precisely at this time point, HOX B6 and EPO gene expression are transferred to both the fetal spleen and fetal kidney. However, surprisingly, expression of both genes increases again in the fetal liver just before birth. HOX B6 is expressed in cells from in vitro erythropoietic colonies (colony-forming unit-erythroid and burst-forming unit-erythroid) and TER-119+ erythroid cells but not in hematopoietic or nonhematopoietic stem cell populations. When the latter two populations are allowed to differentiate into erythropoietic cells, HOX B6 and erythroid-relevant markers are expressed. The results indicate that HOX B6 is intimately involved in the regulation of the erythropoietic system and could be a marker for this lineage.

scholarly journals Cellular and molecular characterization of the role of the flk-2/flt-3 receptor tyrosine kinase in hematopoietic stem cells

Blood ◽  
1994 ◽  
Vol 84 (8) ◽  
pp. 2422-2430 ◽  
Author(s):  
FC Zeigler ◽  
BD Bennett ◽  
CT Jordan ◽  
SD Spencer ◽  
S Baumhueter ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Tyrosine Kinase ◽  
Hematopoietic Stem Cell ◽  
Receptor Tyrosine Kinase ◽  
Lymphoid Cells ◽  
Stem Cell Population ◽  
Cell Populations ◽  
Hematopoietic Stem ◽  
Stem Cell Populations

The flk-2/flt-3 receptor tyrosine kinase was cloned from a hematopoietic stem cell population and is considered to play a potential role in the developmental fate of the stem cell. Using antibodies derived against the extracellular domain of the receptor, we show that stem cells from both murine fetal liver and bone marrow can express flk-2/flt-3. However, in both these tissues, there are stem cell populations that do not express the receptor. Cell cycle analysis shows that stem cells that do not express the receptor have a greater percentage of the population in G0 when compared with the flk-2/flt-3- positive population. Development of agonist antibodies to the receptor shows a proliferative role for the receptor in stem cell populations. Stimulation with an agonist antibody gives rise to an expansion of both myeloid and lymphoid cells and this effect is enhanced by the addition of kit ligand. These studies serve to further illustrate the importance of the flk-2/flt-3 receptor in the regulation of the hematopoietic stem cell.

Leukemic and Normal Stem Cell Transcriptional Signatures Determined by Functional Assays Are Predictive of the Overall Survival of AML Patients.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 389-389
Author(s):  
Kolja Eppert ◽  
Katsuto Takenaka ◽  
Björn Nilsson ◽  
Eric R Lechman ◽  
Vicki Ling ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Overall Survival ◽  
Stem Cell ◽  
Expression Profiles ◽  
Gene Expression Profiles ◽  
Cell Populations ◽  
Hematopoietic Stem ◽  
Functional Assays ◽  
Stem Cell Populations

Abstract Abstract 389 Normal hematopoiesis and acute myeloid leukemia (AML) are organized as hierarchies with stem cells, which possess extensive self-renewal and proliferative capacity, at the apex. Although there is definitive evidence from experimental models for the existence of leukemic stem cells (LSC) in some human leukemias, the relevance of LSC to human disease progression is still lacking. While chemotherapeutic treatment of AML patients typically results in disease remission, the majority of patients will eventually relapse and succumb to the disease, indicating that residual LSC are not eliminated by current treatment. We hypothesize that stem cell derived gene expression profiles may be more clinically relevant than those derived from examination of bulk leukemia samples. Here we show the clinical significance of novel stem cell related expression profiles derived from 25 functionally validated human leukemia stem cell populations and 6 normal hematopoietic stem cell populations. Little is currently known about the molecular regulatory networks that govern human LSC or hematopoietic stem cells (HSC). Therefore, we have carried out global mRNA gene expression profiling of FACS sorted subpopulations of cells enriched for human stem cells, progenitor cells and mature cells from 16 AML primary patient samples and 3 cord blood samples to investigate these pathways. Similar to normal hematopoietic stem cells, leukemia stem, progenitor and mature cells can be sorted using CD34 and CD38 markers. Due to the heterogeneous nature of AML, it is vital that quantitative functional assays are used to characterize the LSC and progenitor activity in each sorted fraction. In vitro cell suspension cultures and methylcellulose colony formation assays were performed to characterize progenitor and blast populations. Importantly, we applied a novel and improved in vivo SCID leukemia initiating cell assay to substantiate the presence of LSC activity in each sorted fraction of 16 AML patient samples. With this enhanced assay, LSC were detected in the expected CD34+/CD38- population. However, in the majority of AML samples, LSC were detected in at least one additional fraction, demonstrating the importance of functional validation when interpreting global gene expression profiles of sorted stem cell populations. LSC and HSC specific signatures were identified following a statistical analysis that compared fractions with stem cell activity against those without (25 LSC vs 29 non-LSC; 6 HSC vs 6 non-HSC). When applied to an independent gene expression data set from 160 cytogenetically normal AML samples, a 25 probe LSC signature was the strongest predictor of overall survival (p<0.0001, HR=2.6, 95%CI 1.8-4.0, median survival 236 vs 999 days; Figure 1a). Furthermore, the 225 probe HSC specific signature derived from normal cells also provided a strong predictor of survival (p<0.0001, HR=2.3, 95%CI 1.5-3.4, median survival 238 vs 741 days; Figure 1b). We queried the gene expression-based chemical genomic database Connectivity Map with the LSC-related gene list and found a negative correlation between the genes in the LSC profile and the expression of genes that are transcriptionally induced following treatment with common chemotherapeutic compounds such as doxorubicin, suggesting resistance to chemotherapy as one possible mechanism for the correlation of the stem cell signatures with survival. Together these data support the hypothesis that the biological determinants that underlie stemness in both normal and leukemic cells are predictors of poor outcome, and are potential targets for novel therapy. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Cellular and molecular characterization of the role of the flk-2/flt-3 receptor tyrosine kinase in hematopoietic stem cells

Blood ◽  
1994 ◽  
Vol 84 (8) ◽  
pp. 2422-2430 ◽  
Author(s):  
FC Zeigler ◽  
BD Bennett ◽  
CT Jordan ◽  
SD Spencer ◽  
S Baumhueter ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Tyrosine Kinase ◽  
Hematopoietic Stem Cell ◽  
Receptor Tyrosine Kinase ◽  
Lymphoid Cells ◽  
Stem Cell Population ◽  
Cell Populations ◽  
Hematopoietic Stem ◽  
Stem Cell Populations

Abstract The flk-2/flt-3 receptor tyrosine kinase was cloned from a hematopoietic stem cell population and is considered to play a potential role in the developmental fate of the stem cell. Using antibodies derived against the extracellular domain of the receptor, we show that stem cells from both murine fetal liver and bone marrow can express flk-2/flt-3. However, in both these tissues, there are stem cell populations that do not express the receptor. Cell cycle analysis shows that stem cells that do not express the receptor have a greater percentage of the population in G0 when compared with the flk-2/flt-3- positive population. Development of agonist antibodies to the receptor shows a proliferative role for the receptor in stem cell populations. Stimulation with an agonist antibody gives rise to an expansion of both myeloid and lymphoid cells and this effect is enhanced by the addition of kit ligand. These studies serve to further illustrate the importance of the flk-2/flt-3 receptor in the regulation of the hematopoietic stem cell.

Leukemia Stem Cell Potential of Different Progenitor Subpopulations in Myeloid Blast Phase CML

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 3489-3489
Author(s):  
Ross Kinstrie ◽  
Dimitris Karamitros ◽  
Nicolas Goardon ◽  
Heather Morrison ◽  
Richard E Clark ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cell ◽  
Research Funding ◽  
Cell Populations ◽  
Novel Therapies ◽  
Blast Phase ◽  
Self Renewal ◽  
Stem Cell Populations

Abstract Blast phase (BP)-CML remains the most critical area of unmet clinical need in the management of CML and novel, targeted therapeutic strategies are urgently needed. In the tyrosine kinase inhibitor (TKI) era, the rate of progression to BP is 1 to 1.5% per annum in the first few years after diagnosis, falling sharply when major molecular response is obtained. Around 10% of patients present with de novo BP-CML and despite the use of TKIs, median survival after the diagnosis of BP-CML is between 6.5 and 11 months.Therefore, improved understanding of the biology of BP-CML and novel therapies to prolong therapeutic responses are urgently sought. Studies of myeloid malignancies show that acquisition of tumor-associated mutations occurs principally in a step-wise manner. Initiating mutations usually originate in an hematopoietic stem cell (HSC) to give rise to preleukemic stem cell populations that expand through clonal advantage. Further mutation acquisition and/or epigenetic changes then lead to blast transformation and disruption of the normal immunophenotypic and functional hematopoietic hierarchy. At this stage, multiple leukemic stem cell (LSC) populations (also termed leukemia initiating cell populations) can be identified. We previously showed, in AML, that the CD34+ LSC populations were most closely related to normal progenitor populations, rather than stem cell populations, but had co-opted elements of a normal stem cell expression signature to acquire abnormal self-renewal potential (Goardon et al, Cancer Cell, 2011). CD34+CD38- LSCs were most commonly similar to an early multi-potent progenitor population with lympho-myeloid potential (the lymphoid-primed multi-potential progenitor [LMPP]). In contrast, the CD34+CD38+ LSCs were most closely related to the more restricted granulocyte-macrophage progenitor (GMP). In chronic phase CML, the leukemia-propagating population is the HSC, and the progenitor subpopulations do not have stem cell characteristics. To date, studies to isolate LSC populations in BP-CML have been limited, identifying the GMP subpopulation only as a possible LSC source (Jamieson et al, NEJM, 2004). Furthermore, in vivo LSC activity has not been assessed. We therefore set out to assess the LSC characteristics of different primitive progenitor subpopulations in myeloid BP-CML both in vitro and in vivo. We isolated different stem and progenitor cell subpopulations using FACS; HSC (Lin-CD34+CD38-CD90+ CD45RA-), multipotent progenitor (MPP; Lin-CD34+CD38-CD90-CD45RA-), LMPP (Lin-CD34+CD38-CD90-CD45RA+), common myeloid progenitor (CMP; Lin-CD34+CD38+CD45RA-CD123+), GMP (Lin-CD34+CD38+CD45RA+CD123+) and megakaryocyte erythroid progenitor (MEP; Lin-CD34+CD38+CD45RA-CD123-). The functional potential of these purified populations was examined in 13 patients by: (i) serial CFC replating assays to study progenitor self-renewal (n=10); (ii) In vivo xenograft studies using NSG mice with serial transplantation to identify populations with LSC potential (n=6). Our data conclusively demonstrate that functional LSCs are present in multiple immunophenotypic stem/progenitor subpopulations in myeloid BP-CML, including HSC, MPP, LMPP, CMP and GMP subpopulations. There was inter-patient variability in terms of both in vitro and in vivo functional properties. Fluorescence in situ hybridisation (FISH) was used to assess clonality in the different progenitor subpopulations and identify which populations contained cells with additional cytogenetic abnormalities (ACAs) with a view to improving our understanding of the clonal hierarchy. Interestingly, there were no significant differences in ACAs in the different progenitor subpopulations in the majority of samples studied, suggesting that clonal evolution tends to occur in the HSC compartment in myeloid BP-CML. Preliminary gene expression profiling studies of the different progenitor subpopulations, using Affymetrix Human Gene 1.0 ST Arrays, demonstrated highly variable gene expression, supporting the functional heterogeneity seen. Taken together, our results demonstrate that myeloid BP-CML is a very heterogeneous disorder with variable LSC populations. Further interrogation of these populations will likely identify novel therapies which will specifically target the LSC. Disclosures Copland: Bristol-Myers Squibb: Consultancy, Honoraria, Other, Research Funding; Novartis: Consultancy, Honoraria, Other; Ariad: Consultancy, Honoraria, Research Funding.

Identification of a Developmentally-Restricted Hematopoietic Stem Cell That Gives Rise to Innate-like Lymphocytes

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4302-4302
Author(s):  
Anna E Beaudin ◽  
Scott W. Boyer ◽  
Gloria Hernandez ◽  
Camilla E Forsberg
Keyword(s):  
Stem Cell ◽  
Hematopoietic Stem Cell ◽  
Conflicts Of Interest ◽  
Fetal Liver ◽  
Lineage Tracing ◽  
Lymphoid Cells ◽  
Gfp Expression ◽  
Hematopoietic Stem

Abstract The generation of innate-like immune cells distinguishes fetal hematopoiesis from adult hematopoiesis, but the cellular mechanisms underlying differential cell production during development remain to be established. Specifically, whether differential lymphoid output arises as a consequence of discrete hematopoietic stem cell (HSC) populations present during development or whether the fetal/neonatal microenvironment is required for their production remains to be established. We recently established a Flk2/Flt3 lineage tracing mouse model wherein Flk2-driven expression of Cre recombinase results in the irreversible switching of a ubiquitous dual-color reporter from Tomato to GFP expression. Because the switch from Tom to GFP expression in this model involves an irreversible genetic excision of the Tomato gene, a GFP+ cell can never give rise to Tom+ progeny. Using this model, we have definitively demonstrated that all functional, adult HSC remain Tomato+ and therefore that all developmental precursors of adult HSC lack a history of Flk2 expression. In contrast, adoptive transfer experiments of Tom+ and GFP+ fetal liver Lin-cKit+Sca1+ (KLS) fractions demonstrated that both Tom+ and GFP+ fetal HSC support serial, long-term multilineage reconstitution (LTR) in irradiated adult recipients. We have therefore identified a novel, developmentally restricted HSC that supports long-term multilineage reconstitution upon transplantation into an adult recipient but does not normally persist into adulthood. Developmentally-restricted GFP+ HSC display greater lymphoid potential, and regenerated both innate-like B-1 lymphocytes and Vg3-expressing T lymphocytes to a greater extent than coexisting Tom+ FL and adult HSC. Interestingly, whereas developmental regulation of fetal-specific B-cell subsets appears to be regulated cell-instrinsically, as fetal HSC generated more innate-like B-cells than adult HSC even within an adult environment, T-cell development may be regulated both cell intrinsically and extrinsically, as both the cell-of-origin and the fetal microenvironment regulated the generation of innate-like T-cells. Our results provide direct evidence for a developmentally restricted HSC that gives rise to a layered immune system and describes a novel mechanism underlying the source of developmental hematopoietic waves. As early lymphoid cells play essential roles in establishing self-recognition and tolerance, these findings are critical for understanding the development of autoimmune diseases, allergies, and tolerance induction upon organ transplantation. Furthermore, by uncoupling self-renewal capacity in situ with that observed upon transplantation, our data suggests that transplantation- and/or irradiation-induced cues may allow for the engraftment of developmental HSC populations that do not normally persist in situ. As LTR upon transplantation has served as the prevailing definition of adult HSC origin during development, our data challenge the current conceptual framework of adult HSC origin. Disclosures No relevant conflicts of interest to declare.

Heterogeneity in hematopoietic stem cell populations

2013 ◽  
Vol 20 (4) ◽  
pp. 257-264 ◽  
Author(s):  
Paul H. Miller ◽  
David J.H.F. Knapp ◽  
Connie J. Eaves
Keyword(s):  
Stem Cell ◽  
Hematopoietic Stem Cell ◽  
Cell Populations ◽  
Hematopoietic Stem ◽  
Stem Cell Populations

271 Embryonic hepatoblasts share patterns of gene expression with other stem cell populations

Hepatology ◽  
2003 ◽  
Vol 38 ◽  
pp. 286-287
Author(s):  
T ADER ◽  
R NOREL ◽  
C ROGLER ◽  
L ROGLER
Keyword(s):  
Gene Expression ◽  
Stem Cell ◽  
Cell Populations ◽  
Stem Cell Populations

Inactivation of a GFP retrovirus occurs at multiple levels in long-term repopulating stem cells and their differentiated progeny

Blood ◽  
2000 ◽  
Vol 96 (3) ◽  
pp. 894-901 ◽  
Author(s):  
Christopher A. Klug ◽  
Samuel Cheshier ◽  
Irving L. Weissman
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Gene Therapy ◽  
Stem Cell ◽  
Viral Vectors ◽  
Fluorescent Protein ◽  
Lymphoid Cells ◽  
Gfp Expression ◽  
Hematopoietic Stem

Abstract Hematopoietic stem cell gene therapy holds promise for the treatment of many hematologic disorders. One major variable that has limited the overall success of gene therapy to date is the lack of sustained gene expression from viral vectors in transduced stem cell populations. To understand the basis for reduced gene expression at a single-cell level, we have used a murine retroviral vector, MFG, that expresses the green fluorescent protein (GFP) to transduce purified populations of long-term self-renewing hematopoietic stem cells (LT-HSC) isolated using the fluorescence-activated cell sorter. Limiting dilution reconstitution of lethally irradiated recipient mice with 100% transduced, GFP+ LT-HSC showed that silencing of gene expression occurred rapidly in most integration events at the LT-HSC level, irrespective of the initial levels of GFP expression. When inactivation occurred at the LT-HSC level, there was no GFP expression in any hematopoietic lineage clonally derived from silenced LT-HSC. Inactivation downstream of LT-HSC that stably expressed GFPin long-term reconstituted animals was restricted primarily to lymphoid cells. These observations suggest at least 2 distinct mechanisms of silencing retrovirally expressed genes in hematopoietic cells.

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