N-cadherin identifies human endometrial epithelial progenitor cells by in vitro stem cell assays

Abstract Stem cell factor (SCF) and erythropoietin (Epo) effectively support erythroid cell development in vivo and in vitro. We have studied here an SCF/Epo-dependent erythroid progenitor cell from cord blood that can be efficiently amplified in liquid culture to large cell numbers in the presence of SCF, Epo, insulin-like growth factor-1 (IGF-1), dexamethasone, and estrogen. Additionally, by changing the culture conditions and by administration of Epo plus insulin, such progenitor cells effectively undergo terminal differentiation in culture and thereby faithfully recapitulate erythroid cell differentiation in vitro. This SCF/Epo-dependent erythroid progenitor is also present in CD34+ peripheral blood stem cells and human bone marrow and can be isolated, amplified, and differentiated in vitro under the same conditions. Thus, highly homogenous populations of SCF/Epo-dependent erythroid progenitors can be obtained in large cell numbers that are most suitable for further biochemical and molecular studies. We demonstrate that such cells express the recently identified adapter protein p62dok that is involved in signaling downstream of the c-kit/SCF receptor. Additionally, cells express the cyclin-dependent kinase (CDK) inhibitors p21cip1 and p27kip1 that are highly induced when cells differentiate. Thus, the in vitro system described allows the study of molecules and signaling pathways involved in proliferation or differentiation of human erythroid cells.

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Hypoxia Enhances the Expansion of Human Limbal Epithelial Progenitor Cells In Vitro

Investigative Opthalmology & Visual Science ◽

10.1167/iovs.07-0077 ◽

2007 ◽

Vol 48 (8) ◽

pp. 3586 ◽

Cited By ~ 23

Author(s):

Hideyuki Miyashita ◽

Kazunari Higa ◽

Naoko Kato ◽

Tetsuya Kawakita ◽

Satoru Yoshida ◽

...

Keyword(s):

Progenitor Cells ◽

Epithelial Progenitor Cells

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Heparanase regulates retention and proliferation of primitive Sca-1+/c-Kit+/Lin− cells via modulation of the bone marrow microenvironment

Blood ◽

10.1182/blood-2007-10-116145 ◽

2008 ◽

Vol 111 (10) ◽

pp. 4934-4943 ◽

Cited By ~ 27

Author(s):

Asaf Spiegel ◽

Eyal Zcharia ◽

Yaron Vagima ◽

Tomer Itkin ◽

Alexander Kalinkovich ◽

...

Keyword(s):

Bone Marrow ◽

Stem Cell ◽

Progenitor Cells ◽

Progenitor Cell ◽

Cathepsin K ◽

Hematopoietic Progenitors ◽

Hematopoietic Progenitor ◽

Cell Mobilization ◽

Tumor Growth And Metastasis

Abstract Heparanase is involved in tumor growth and metastasis. Because of its unique cleavage of heparan sulfate, which binds cytokines, chemokines and proteases, we hypothesized that heparanase is also involved in regulation of early stages of hematopoiesis. We report reduced numbers of maturing leukocytes but elevated levels of undifferentiated Sca-1+/c-Kit+/Lin− cells in the bone marrow (BM) of mice overexpressing heparanase (hpa-Tg). This resulted from increased proliferation and retention of the primitive cells in the BM microenvironment, manifested in increased SDF-1 turnover. Furthermore, heparanase overexpression in mice was accompanied by reduced protease activity of MMP-9, elastase, and cathepsin K, which regulate stem and progenitor cell mobilization. Moreover, increased retention of the progenitor cells also resulted from up-regulated levels of stem cell factor (SCF) in the BM, in particular in the stem cell–rich endosteum and endothelial regions. Increased SCF-induced adhesion of primitive Sca-1+/c-Kit+/Lin− cells to osteoblasts was also the result of elevation of the receptor c-Kit. Regulation of these phenomena is mediated by hyperphosphorylation of c-Myc in hematopoietic progenitors of hpa-Tg mice or after exogenous heparanase addition to wildtype BM cells in vitro. Altogether, our data suggest that heparanase modification of the BM microenvironment regulates the retention and proliferation of hematopoietic progenitor cells.

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N-Cadherin in the Maintenance of Human Corneal Limbal Epithelial Progenitor Cells In Vitro

Investigative Opthalmology & Visual Science ◽

10.1167/iovs.09-3503 ◽

2009 ◽

Vol 50 (10) ◽

pp. 4640 ◽

Cited By ~ 22

Author(s):

Kazunari Higa ◽

Shigeto Shimmura ◽

Hideyuki Miyashita ◽

Naoko Kato ◽

Yoko Ogawa ◽

...

Keyword(s):

Progenitor Cells ◽

Epithelial Progenitor Cells

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Chemotactic and chemokinetic activities of stem cell factor on murine hematopoietic progenitor cells

Blood ◽

10.1182/blood.v87.10.4100.bloodjournal87104100 ◽

1996 ◽

Vol 87 (10) ◽

pp. 4100-4108 ◽

Cited By ~ 61

Author(s):

N Okumura ◽

K Tsuji ◽

Y Ebihara ◽

I Tanaka ◽

N Sawai ◽

...

Keyword(s):

Bone Marrow ◽

Stem Cell ◽

Progenitor Cells ◽

Stromal Cells ◽

Stem Cell Factor ◽

Hematopoietic Progenitor Cells ◽

Hematopoietic Progenitor ◽

Murine Bone Marrow ◽

Checkerboard Assay

We investigated the effects of stem cell factor (SCF) on the migration of murine bone marrow hematopoietic progenitor cells (HPC) in vitro using a modification of the checkerboard assay. Chemotactic and chemokinetic activities of SCF on HPC were evaluated by the numbers of HPC migrated on positive and negative gradients of SCF, respectively. On both positive and negative gradients of SCF, HPC began to migrate after 4 hours incubation, and their numbers then increased time- dependently. These results indicated that SCF functions as a chemotactic and chemokinetic agent for HPC. Analysis of types of colonies derived from the migrated HPC showed that SCF had chemotactic and chemokinetic effects on all types of HPC. When migrating activities of other cytokines were examined, interleukin (IL)-3 and IL-11 also affected the migration of HPC, but the degrees of each effect were lower than that of SCF. The results of the present study demonstrated that SCF is one of the most potent chemotactic and chemokinetic factors for HPC and suggest that SCF may play an important role in the flow of HPC into bone marrow where stromal cells constitutively produce SCF.

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Human Cytomegalovirus IE2 Protein Disturbs Brain Development by the Dysregulation of Neural Stem Cell Maintenance and the Polarization of Migrating Neurons

Journal of Virology ◽

10.1128/jvi.00799-17 ◽

2017 ◽

Vol 91 (17) ◽

Cited By ~ 6

Author(s):

Dasol Han ◽

Sung-Hyun Byun ◽

Juwan Kim ◽

Mookwang Kwon ◽

Samuel J. Pleasure ◽

...

Keyword(s):

Stem Cell ◽

Brain Development ◽

Progenitor Cells ◽

Neural Stem Cell ◽

Human Cytomegalovirus ◽

Neural Progenitor Cells ◽

Hcmv Infection ◽

Stem Cell Maintenance ◽

Cell Maintenance

ABSTRACT Despite the high incidence of severe defects in the central nervous system caused by human cytomegalovirus (HCMV) congenital infection, the mechanism of HCMV neuropathogenesis and the roles of individual viral genes have not yet been fully determined. In this study, we show that the immediate-early 2 (IE2) protein may play a key role in HCMV-caused neurodevelopmental disorders. IE2-transduced neural progenitor cells gave rise to neurospheres with a lower frequency and produced smaller neurospheres than control cells in vitro, indicating reduction of self-renewal and expansion of neural progenitors by IE2. At 2 days after in utero electroporation into the ventricle of the developing brain, a dramatically lower percentage of IE2-expressing cells was detected in the ventricular zone (VZ) and cortical plate (CP) compared to control cells, suggesting that IE2 concurrently dysregulates neural stem cell maintenance in the VZ and neuronal migration to the CP. In addition, most IE2+ cells in the lower intermediate zone either showed multipolar morphology with short neurites or possessed nonradially oriented processes, whereas control cells had long, radially oriented monopolar or bipolar neurites. IE2+ callosal axons also failed to cross the midline to form the corpus callosum. Furthermore, we provide molecular evidence that the cell cycle arrest and DNA binding activities of IE2 appear to be responsible for the increased neural stem cell exit from the VZ and cortical migrational defects, respectively. Collectively, our results demonstrate that IE2 disrupts the orderly process of brain development in a stepwise manner to further our understanding of neurodevelopmental HCMV pathogenesis. IMPORTANCE HCMV brain pathogenesis has been studied in limited experimental settings, such as in vitro HCMV infection of neural progenitor cells or in vivo murine CMV infection of the mouse brain. Here, we show that IE2 is a pivotal factor that contributes to HCMV-induced abnormalities in the context of the embryonic brain using an in utero gene transfer tool. Surprisingly, IE2, but not HCMV IE1 or murine CMV ie3, interferes pleiotropically with key neurodevelopmental processes, including neural stem cell regulation, proper positioning of migrating neurons, and the callosal axon projections important for communication between the hemispheres. Our data suggest that the wide spectrum of clinical outcomes, ranging from mental retardation to microcephaly, caused by congenital HCMV infection can be sufficiently explained in terms of IE2 action alone.

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Involvement of the Integrin Alpha 6 Receptor in the Homing and Engraftment of Hematopoietic Stem and Progenitor Cells to Bone Marrow.

Blood ◽

10.1182/blood.v104.11.1293.1293 ◽

2004 ◽

Vol 104 (11) ◽

pp. 1293-1293

Author(s):

Hong Qian ◽

Sten Eirik W. Jacobsen ◽

Marja Ekblom

Keyword(s):

Bone Marrow ◽

Stem Cell ◽

Progenitor Cells ◽

Progenitor Cell ◽

Bone Marrow Cells ◽

Hematopoietic Stem ◽

Stem And Progenitor Cells ◽

Functional Blocking

Abstract Within the bone marrow environment, adhesive interactions between stromal cells and extracellular matrix molecules are required for stem and progenitor cell survival, proliferation and differentiation as well as their transmigration between bone marrow (BM) and the circulation. This regulation is mediated by cell surface adhesion receptors. In experimental mouse stem cell transplantation models, several classes of cell adhesion receptors have been shown to be involved in the homing and engraftment of stem and progenitor cells in BM. We have previously found that integrin a6 mediates human hematopoietic stem and progenitor cell adhesion to and migration on its specific ligands, laminin-8 and laminin-10/11 in vitro (Gu et al, Blood, 2003; 101:877). Using FACS analysis, the integrin a6 chain was now found to be ubiquitously (>95%) expressed in mouse hematopoietic stem and progenitor cells (lin−Sca-1+c-Kit+, lin−Sca-1+c-Kit+CD34+) both in adult bone marrow and in fetal liver. In vitro, about 70% of mouse BM lin−Sca-1+c-Kit+ cells adhered to laminin-10/11 and 40% adhered to laminin-8. This adhesion was mediated by integrin a6b1 receptor, as shown by functional blocking monoclonal antibodies. We also used a functional blocking monoclonal antibody (GoH3) against integrin a6 to analyse the role of the integrin a6 receptor for the in vivo homing of hematopoietic stem and progenitor cells. We found that the integrin a6 antibody inhibited the homing of bone marrow progenitors (CFU-C) into BM of lethally irradiated recipients. The number of homed CFU-C was reduced by about 40% as compared to cells incubated with an isotype matched control antibody. To study homing of long-term repopulating stem cells (LTR), antibody treated bone marrow cells were first injected intravenously into lethally irradiated primary recipients. After three hours, bone marrow cells of the primary recipients were analysed by competitive repopulation assay in secondary recipients. Blood analysis 16 weeks after transplantation revealed an 80% reduction of stem cell activity of integrin a6 antibody treated cells as compared to cells treated with control antibody. These results suggest that integrin a6 plays an important role for hematopoietic stem and progenitor cell homing in vivo.

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