Successful elimination of non-neural cells and unachievable elimination of glial cells by means of commonly used cell culture manipulations during differentiation of GFAP and SOX2 positive neural progenitors (NHA) to neuronal cells

Witusik, Monika; Piaskowski, Sylwester; Hułas-Bigoszewska, Krystyna; Zakrzewska, Magdalena; Gresner, Sylwia M.; Azizi, S. Ausim; Krynska, Barbara; Liberski, Paweł P.; Rieske, Piotr

doi:10.1186/1472-6750-8-56

Cited by 13 publications

(16 citation statements)

References 27 publications

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“…Among cells showing GFAP and MAP2 co-expression, most (about 70%) presented only the remnants of GFAP, which constitutes additional evidence that the neuronal cells originated from the cells initially showing a multilineage phenotype. The existence of MAP2+, GFAP+, CD44- cells has already been presented by our group [7]. We showed that the percentage of those cells after 5–7 days of neural differentiation did not exceed 5% [7].…”

Section: Resultssupporting

confidence: 55%

“…The existence of MAP2+, GFAP+, CD44- cells has already been presented by our group [7]. We showed that the percentage of those cells after 5–7 days of neural differentiation did not exceed 5% [7]. …”

Section: Resultssupporting

confidence: 55%

“…All undifferentiated GFAP+NNP presented the multilineage phenotype, defined as co-expression of GFAP, CD44, Beta III-tubulin, MAP2 and Nestin, SOX-2, Vimentin [5-7]. GFAP+NNP were CD133 negative (Fig.…”

Section: Resultsmentioning

confidence: 99%

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Arrested neural and advanced mesenchymal differentiation of glioblastoma cells-comparative study with neural progenitors

et al. 2009

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BackgroundAlthough features of variable differentiation in glioblastoma cell cultures have been reported, a comparative analysis of differentiation properties of normal neural GFAP positive progenitors, and those shown by glioblastoma cells, has not been performed.MethodsFollowing methods were used to compare glioblastoma cells and GFAP+NNP (NHA): exposure to neural differentiation medium, exposure to adipogenic and osteogenic medium, western blot analysis, immunocytochemistry, single cell assay, BrdU incorporation assay. To characterize glioblastoma cells EGFR amplification analysis, LOH/MSI analysis, and P53 nucleotide sequence analysis were performed.ResultsIn vitro differentiation of cancer cells derived from eight glioblastomas was compared with GFAP-positive normal neural progenitors (GFAP+NNP). Prior to exposure to differentiation medium, both types of cells showed similar multilineage phenotype (CD44+/MAP2+/GFAP+/Vimentin+/Beta III-tubulin+/Fibronectin+) and were positive for SOX-2 and Nestin. In contrast to GFAP+NNP, an efficient differentiation arrest was observed in all cell lines isolated from glioblastomas. Nevertheless, a subpopulation of cells isolated from four glioblastomas differentiated after serum-starvation with varying efficiency into derivatives indistinguishable from the neural derivatives of GFAP+NNP. Moreover, the cells derived from a majority of glioblastomas (7 out of 8), as well as GFAP+NNP, showed features of mesenchymal differentiation when exposed to medium with serum.ConclusionOur results showed that stable co-expression of multilineage markers by glioblastoma cells resulted from differentiation arrest. According to our data up to 95% of glioblastoma cells can present in vitro multilineage phenotype. The mesenchymal differentiation of glioblastoma cells is advanced and similar to mesenchymal differentiation of normal neural progenitors GFAP+NNP.

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Section: Resultssupporting

confidence: 55%

Section: Resultssupporting

confidence: 55%

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Arrested neural and advanced mesenchymal differentiation of glioblastoma cells-comparative study with neural progenitors

et al. 2009

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“…2A and B). The NPs also expressed GFAP, an astrocyte and neural progenitor cell marker (Schwartz et al, 2003;Witusik et al, 2008). In addition, immunoreactivity was detected for Oct-4, a marker for hESC pluripotency, although the gel electrophoresis product band was weak.…”

Section: Genementioning

confidence: 96%

A method for rapid derivation and propagation of neural progenitors from human embryonic stem cells

Axell

Zlateva

Curtis

2009

Journal of Neuroscience Methods

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“…This consequently could have resulted in a value more than 100% for the combined percentages of cells that stained positive for the examined markers in GS‐2 and D3 cell lines. In this context, previous studies, based on the culture of brain‐derived cells, have also shown coexpression of these neural markers in neural progenitors (Witusik et al, 2008), astrocytes (Dráberová et al, 2008), and brain parenchymal cells (Rieske et al, 2007).…”

Section: Discussionmentioning

confidence: 81%

Efficient neural differentiation of mouse pluripotent stem cells in a serum‐free medium and development of a novel strategy for enrichment of neural cells

Verma

Rashid

Sikdar

et al. 2017

Intl J of Devlp Neuroscience

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Pluripotent stem cells (PSCs) offer an excellent model to study neural development and function. Although various protocols have been developed to direct the differentiation of PSCs into desired neural cell types, many of them suffer from limitations including low efficiency, long duration of culture, and the use of expensive, labile, and undefined growth supplements. In this study, we achieved efficient differentiation of mouse PSCs to neural lineage, in the absence of exogenous molecules, by employing a serum-free culture medium containing knockout serum replacement (KSR). Embryoid bodies (EBs) cultured in this medium predominantly produced neural cells which included neural progenitors (15-18%), immature neurons (8-24%), mature neurons (10-26%), astrocytes (27-61%), and oligodendrocytes (∼1%). Different neuronal subtypes including glutamatergic, GABAergic, cholinergic, serotonergic, and dopaminergic neurons were generated. Importantly, neurons generated in the KSR medium were electrically active. Further, the EB scooping strategy, involving the removal of the EB core region from the peripheral EB outgrowth, resulted in the enrichment of PSC-derived neural cells. Taken together, this study provides the evidence that the KSR medium is ideal for the rapid and efficient generation of neural cells, including functional neurons, from PSCs without the requirement of any other additional molecule.

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Successful elimination of non-neural cells and unachievable elimination of glial cells by means of commonly used cell culture manipulations during differentiation of GFAP and SOX2 positive neural progenitors (NHA) to neuronal cells

Cited by 13 publications

References 27 publications

Arrested neural and advanced mesenchymal differentiation of glioblastoma cells-comparative study with neural progenitors

Arrested neural and advanced mesenchymal differentiation of glioblastoma cells-comparative study with neural progenitors

A method for rapid derivation and propagation of neural progenitors from human embryonic stem cells

Efficient neural differentiation of mouse pluripotent stem cells in a serum‐free medium and development of a novel strategy for enrichment of neural cells

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