Physiologic Oxygen Concentration Enhances the Stem-Like Properties of CD133(+) Human Glioblastoma Cells In vitro

Moffitt Cancer Center, Tampa, FL 33612, USA.
Molecular Cancer Research (Impact Factor: 4.5). 05/2009; 7(4):489-97. DOI: 10.1158/1541-7786.MCR-08-0360
Source: PubMed

ABSTRACT In vitro investigations of tumor stem-like cells (TSC) isolated from human glioblastoma (GB) surgical specimens have been done primarily at an atmospheric oxygen level of 20%. To determine whether an oxygen level more consistent with in situ conditions affects their stem cell-like characteristics, we compared GB TSCs grown under conditions of 20% and 7% oxygen. Growing CD133(+) cells sorted from three GB neurosphere cultures at 7% O(2) reduced their doubling time and increased the self-renewal potential as reflected by clonogenicity. Furthermore, at 7% oxygen, the cultures exhibited an enhanced capacity to differentiate along both the glial and neuronal pathways. As compared with 20%, growth at 7% oxygen resulted in an increase in the expression levels of the neural stem cell markers CD133 and nestin as well as the stem cell markers Oct4 and Sox2. In addition, whereas hypoxia inducible factor 1alpha was not affected in CD133(+) TSCs grown at 7% O(2), hypoxia-inducible factor 2alpha was expressed at higher levels as compared with 20% oxygen. Gene expression profiles generated by microarray analysis revealed that reducing oxygen level to 7% resulted in the up-regulation and down-regulation of a significant number of genes, with more than 140 being commonly affected among the three CD133(+) cultures. Furthermore, Gene Ontology categories up-regulated at 7% oxygen included those associated with stem cells or GB TSCs. Thus, the data presented indicate that growth at the more physiologically relevant oxygen level of 7% enhances the stem cell-like phenotype of CD133(+) GB cells.

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Available from: Muhammad Jamal, Aug 25, 2015
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    • "We assessed the percentages of CD133 positivity, a surrogate marker for GSCs, present in three freshly disaggregated xenolines (GBM- XD456, GBM-X12, and GBM-X6) in tissue culture under normoxic and 1% hypoxic environments. Similar to other reports, CD133 expression increased significantly in all three xenolines under hypoxia (Table 1) [8] [9]. GBM-XD456, which had the lowest percentage of CD133 + cells in normoxia (16.5%), had the greatest relative increase in CD133 expression (nearly four-fold) under hypoxia. "
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    • "Neurospheres were maintained in medium consisting of Dulbecco's modified Eagle medium (DMEM)/F-12 (Invitrogen, Carlsbad, CA), B27 supplement (1×; Invitrogen), and human recombinant basic fibroblast growth factor and epidermal growth factor (50 ng/ml each; R&D Systems, Minneapolis, MN). All cultures were maintained at 37°C in an atmosphere of 5% CO 2 /7% O 2 [19]. CD133 + cells were isolated from each neurosphere cultures by FACS [10] and used as a source for the described experiments. "
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