Derivation and spontaneous differentiation of human embryonic stem cells*

Amit, Michal; Itskovitz‐Eldor, Joseph

doi:10.1046/j.1469-7580.2002.00032.x

Cited by 198 publications

(102 citation statements)

References 37 publications

(72 reference statements)

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…HESC lines I3, I4 19 and H1 4 were cultured with inactivated MEF (mouse embryonic fibroblasts) as was previously described 11 for 19-102 passages.…”

Section: Culturing Hesc and Hipsc Linesmentioning

confidence: 99%

Screening of human pluripotent stem cells using CGH and FISH reveals low-grade mosaic aneuploidy and a recurrent amplification of chromosome 1q

et al. 2012

Self Cite

View full text Add to dashboard Cite

Pluripotency and proliferative capacity of human embryonic stem cells (hESCs) make them a promising source for basic and applied research as well as in therapeutic medicine. The introduction of human induced pluripotent cells (hiPSCs) holds great promise for patient-tailored regenerative medicine therapies. However, for hESCs and hiPSCs to be applied for therapeutic purposes, long-term genomic stability in culture must be maintained. Until recently, G-banding analysis was considered as the default approach for detecting chromosomal abnormalities in stem cells. Our goal in this study was to apply fluorescence in-situ hybridization (FISH) and comparative genomic hybridization (CGH) for the screening of pluripotent stem cells, which will enable us identifying chromosomal abnormalities in stem cells genome with a better resolution. We studied three hESC lines and two hiPSC lines over long-term culture. Aneuploidy rates were evaluated at different passages, using FISH probes (12,13,16,17,18,21,X,Y). Genomic integrity was shown to be maintained at early passages of hESCs and hiPSCs but, at late passages, we observed low rates mosaiciam in hESCs, which implies a direct correlation between number of passages and increased aneuploidy rate. In addition, CGH analysis revealed a recurrent genomic instability, involving the gain of chromosome 1q. This finding was detected in two unrelated cell lines of different origin and implies that gains of chromosome 1q may endow a clonal advantage in culture. These findings, which could only partially be detected by conventional cytogenetic methods, emphasize the importance of using molecular cytogenetic methods for tracking genomic instability in stem cells.

show abstract

“…HESC lines I3, I4 19 and H1 4 were cultured with inactivated MEF (mouse embryonic fibroblasts) as was previously described 11 for 19-102 passages.…”

Section: Culturing Hesc and Hipsc Linesmentioning

confidence: 99%

Screening of human pluripotent stem cells using CGH and FISH reveals low-grade mosaic aneuploidy and a recurrent amplification of chromosome 1q

et al. 2012

Self Cite

View full text Add to dashboard Cite

show abstract

“…Approximately 20 lines from the 78 derivations undertaken before August 9, 2001, are available in sufficient quantities for general research use (National Institutes of Health [NIH] stem cell registry, http://stemcells.nih.gov/research/registry). Of these, only a small subset of lines is available for detailed characterization [1][2][3][4][5][6][7][8]. As expected, various hESC lines have a number of similarities.…”

Section: Introductionmentioning

confidence: 99%

“…As expected, various hESC lines have a number of similarities. For example, undifferentiated hESCs are similar in expressing surface antigens and markers characteristic of the undifferentiated ESC state, including Oct4 (POU5F1), Nanog, UTF1, DPPA5, TERT, gap junction proteins, and SSEA and TRA antigens [1][2][3][4][5][6][7][8]. hESCs are also similar in their ability to proliferate and differentiate into cell types of the three germ layers in vitro and in vivo [9 -16].…”

Section: Introductionmentioning

confidence: 99%

Assessing Self-Renewal and Differentiation in Human Embryonic Stem Cell Lines

et al. 2005

View full text Add to dashboard Cite

Like other cell populations, undifferentiated human embryonic stem cells (hESCs) express a characteristic set of proteins and mRNA that is unique to the cells regardless of culture conditions, number of passages, and methods of propagation. We sought to identify a small set of markers that would serve as a reliable indicator of the balance of undifferentiated and differentiated cells in hESC populations. Markers of undifferentiated cells should be rapidly downregulated as the cells differentiate to form embryoid bodies (EBs), whereas markers that are absent or low during the undifferentiated state but that are induced as hESCs differentiate could be used to assess the presence of differentiated cells in the cultures. In this paper, we describe a list of markers that reliably distinguish undifferentiated and differentiated cells. An initial list of approximately 150 genes was generated by scanning published massively parallel signature sequencing, expressed sequence tag scan, and microarray datasets. From this list, a subset of 109 genes was selected that included 55 candidate markers of undifferentiated cells, 46 markers of hESC derivatives, four germ cell markers, and four trophoblast markers. Expression of these candidate marker genes was analyzed in undifferentiated hESCs and differentiating EB populations in four different lines by immunocytochemistry, reverse transcription-polymerase chain reaction (RT-PCR), microarray analysis, and quantitative RT-PCR (qPCR). We show that qPCR, with as few as 12 selected genes, can reliably distinguish differentiated cells from undifferentiated hESC populations.

show abstract

“…hES cells are generally derived from the inner cell mass of blastocysts produced by in vitro fertilization techniques (1,3,(10)(11)(12)(13). It is not widely appreciated that the environment of the mammalian reproductive tract, to which naturally conceived embryos are exposed, is 1.5-5.3% O 2 (14).…”

mentioning

confidence: 99%

Low O ₂ tensions and the prevention of differentiation of hES cells

Ezashi

Das²,

Roberts³

2005

Proc. Natl. Acad. Sci. U.S.A.

756

600

View full text Add to dashboard Cite

Early-stage mammalian embryos develop in a low O2 environment (hypoxia). hES cells, however, are generally cultured under an atmosphere of 21% O2 (normoxia), under which conditions they tend to differentiate spontaneously. Such conditions may not be the most suitable, therefore, for hES cell propagation. Here we have tested two hypotheses. The first hypothesis was that hES cells would grow as well under hypoxic as under normoxic conditions. The second hypothesis was that hypoxic culture would reduce the amount of spontaneous cell differentiation that occurs in hES colonies. Both hypotheses proved to be correct. Cells proliferated as well under 3% and 5% O2 as they did under 21% O2, and growth was only slightly reduced at 1% O2. The appearance of differentiated regions as assessed morphologically, biochemically (by the production of human chorionic gonadotropin and progesterone), and immunohistochemically (by the loss of stage-specific embryonic antigen-4 and Oct-4 and gain of stage-specific embryonic antigen-1 marker expression) was markedly reduced under hypoxic conditions. In addition, hES cell growth under hypoxia provided enhanced formation of embryoid bodies. Hypoxic culture would appear to be necessary to maintain full pluripotency of hES cells.H1 cell ͉ hypoxia ͉ pluripotent ͉ human chorionic gonadotropin ͉ Oct-4 transcription factor

show abstract

Derivation and spontaneous differentiation of human embryonic stem cells*

Cited by 198 publications

References 37 publications

Screening of human pluripotent stem cells using CGH and FISH reveals low-grade mosaic aneuploidy and a recurrent amplification of chromosome 1q

Screening of human pluripotent stem cells using CGH and FISH reveals low-grade mosaic aneuploidy and a recurrent amplification of chromosome 1q

Assessing Self-Renewal and Differentiation in Human Embryonic Stem Cell Lines

Low O ₂ tensions and the prevention of differentiation of hES cells

Contact Info

Product

Resources

About

Derivation and spontaneous differentiation of human embryonic stem cells*

Cited by 198 publications

References 37 publications

Screening of human pluripotent stem cells using CGH and FISH reveals low-grade mosaic aneuploidy and a recurrent amplification of chromosome 1q

Screening of human pluripotent stem cells using CGH and FISH reveals low-grade mosaic aneuploidy and a recurrent amplification of chromosome 1q

Assessing Self-Renewal and Differentiation in Human Embryonic Stem Cell Lines

Low O 2 tensions and the prevention of differentiation of hES cells

Contact Info

Product

Resources

About

Low O ₂ tensions and the prevention of differentiation of hES cells