Role of serum on cryopreservation and subsequent viability of mouse bone marrow hemopoietic stem cells

Cryobiology ◽  
1980 ◽  
Vol 17 (5) ◽  
pp. 516-520 ◽  
Author(s):  
G. Grilli ◽  
A. Porcellini ◽  
G. Lucarelli
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mouse Bone Marrow ◽  
Hemopoietic Stem Cells

scholarly journals Isolation of murine pluripotent hemopoietic stem cells.

1984 ◽  
Vol 159 (6) ◽  
pp. 1576-1590 ◽  
Author(s):  
J W Visser ◽  
J G Bauman ◽  
A H Mulder ◽  
J F Eliason ◽  
A M de Leeuw
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Bone Marrow Cells ◽  
Indirect Evidence ◽  
Fluorescein Isothiocyanate ◽  
Cell Types ◽  
Mouse Bone Marrow ◽  
Hemopoietic Stem Cells ◽  
Morphological Criteria

A method described to purify pluripotent hemopoietic stem cells ( PHSC ) from adult mouse bone marrow. The method consists of three separation steps. First, bone marrow cells are centrifuged in a discontinuous metrizamide gradient and simultaneously labeled with wheat germ agglutinin-fluorescein isothiocyanate (WGA-FITC). Second, the low density cells are analyzed by a fluorescence-activated cell sorter (FACS) and the WGA-positive cells with medium forward and low perpendicular light scatter intensities are sorted. The WGA-FITC is removed from the cells by incubation with N-acetyl-D-glucosamine. Finally, the sorted cells are incubated with anti-H-2K-biotin and avidin-FITC and sorted a second time to enrich cells with high H-2K density. The sorted cells gave rise to 2 spleen colonies per 100 injected cells at 8 d and 6.6 colonies per 100 cells at 12 d after transplantation into lethally irradiated syngeneic recipients. The average enrichment factor for day 12 CFU-S (colony-forming unit/spleen) was 135 (range, 90--230; n = 15) and was similar to that for the cell type that provides radioprotection (180 +/- 70), indicating that these functional properties were copurified. Indirect evidence suggests that the spleen-seeding efficiency (f factor) of these cells is 0.10 and, therefore, the average purity of the sorted PHSC was 65% (range in 15 experiments, 35--110%). The sorted cells were all in the G1 or G0 phase of the cell cycle. They appeared to be undifferentiated blasts by morphological criteria. Electron microscopy revealed that the sorted cells consisted primarily of two cell types, possibly representing G0 and G1 cells. The FACS was used to deposit single selected cells into individual microwells of Terasaki trays. 32% of the sorted cells could be induced to form myeloid progeny in vitro. This procedure should be useful for direct studies on the regulation of hemopoietic cell differentiation.

Phenotypically identical hemopoietic stem cells isolated from different regions of bone marrow have different biologic potential

Blood ◽  
2010 ◽  
Vol 116 (17) ◽  
pp. 3185-3196 ◽  
Author(s):  
Jochen Grassinger ◽  
David N. Haylock ◽  
Brenda Williams ◽  
Gemma H. Olsen ◽  
Susan K. Nilsson
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Hemopoietic Stem Cells ◽  
Proliferative Capacity ◽  
Functional Differences ◽  
Hsc Niche ◽  
And Migration ◽  
B Lymphoid ◽  
First Time

Abstract Hemopoietic stem cells (HSCs) reside within a specified area of the bone marrow (BM) cavity called a “niche” that modulates HSC quiescence, proliferation, differentiation, and migration. Our previous studies have identified the endosteal BM region as the site for the HSC niche and demonstrated that hemopoietic stem and progenitor populations (HSPCs, LSK) isolated from different BM regions exhibit significantly different hemopoietic potential. In this study, we have analyzed subpopulations of LSK cells isolated from different regions of the BM and showed that CD150+CD48−LSK HSCs within the endosteal BM region have superior proliferative capacity and homing efficiency compared with CD150+CD48−LSK HSCs isolated from the central BM. Furthermore, we show, for the first time, that a subset of CD150+CD48+LSK progenitor cells, previously defined as B-lymphoid primed hemopoietic cells, are capable of multilineage reconstitution, however, only when isolated from the endosteal region. In addition, we provide evidence for an unrecognized role of CD48 in HSC homing. Together, our data provide strong evidence that highly purified HSCs show functional differences depending on their origin within the BM and that the most primitive HSCs reside within the endosteal BM region.

Nucleotides metabolizing ectoenzymes as possible markers of mesenchymal stem cell osteogenic differentiation

2013 ◽  
Vol 91 (3) ◽  
pp. 176-181 ◽  
Author(s):  
Katarzyna Roszek ◽  
Agnieszka Błaszczak ◽  
Magdalena Wujak ◽  
Michał Komoszyński
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Adenylate Kinase ◽  
Atp Hydrolysis ◽  
Inhibitory Effect ◽  
Nucleoside Triphosphate ◽  
Mouse Bone Marrow ◽  
Nucleoside Triphosphate Diphosphohydrolase ◽  
High Expression Level

Growing murine mesenchymal stem cells (mMSCs) from mouse bone marrow decreased their rate of proliferation in the presence of benzoylbenzoyl-ATP persistently, but the inhibitory effect of ATP was strong only in a concentration of 50 μmol·L−1 and lasted for 48 h in culture. These results hinted at ATP hydrolysis by the cell surface enzymes at the lower concentrations and thus it may be not able to inhibit MSCs. By using ATP, ADP, or AMP as substrates, we tested the ectonucleotidase activity on the surface of undifferentiated MSCs and MSC-derived osteoblasts. Here, we report that although nucleoside triphosphate diphosphohydrolase (NTPDase)1 and NTPDase8 are engaged in the metabolism of ATP in MSC-derived osteoblasts, NTPDase3 is responsible for its metabolism in undifferentiated MSCs. In this study, we also realized that osteoblasts effectively metabolize ADP to ATP and AMP. The enzymatic activity of adenylate kinase (AK) is consistent with the high expression level of the AK gene. Therefore, it was tempting to suggest that this enzyme, together with NTPDase1 and NTPDase8, assume the role of specific markers that allowed distinction between differentiated osteoblasts and early undifferentiated MSCs. Additionally, unlike osteoblasts, undifferentiated MSCs demonstrated the activity of 5′-nucleotidase (CD73). However, the expression analysis of CD73 mRNA did not show any differences; CD73 mRNA was expressed in both kinds of cells to the same extent.

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