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

Cai, Jun; Chen, Jia; Liu, Ying; Miura, Takumi; Luo, Yongquan; Loring, Jeanne F.; Freed, William J.; Rao, Mahendra S.; Zeng, Xianmin

doi:10.1634/stemcells.2005-0143

Cited by 127 publications

(121 citation statements)

References 110 publications

Supporting

Mentioning

115

Contrasting

Order By: Relevance

“…ABI prism 7500 and the analysis was performed similar to that described for quantitative ChIP PCR above, except that DC t was obtained after normalization with C t of Gapdh for mouse-specific primers and that of actin for human-specific primers. Mouse-specific primers will be provided on request; human-specific primers are as published 24 .…”

Section: Methods Summarymentioning

confidence: 99%

REST maintains self-renewal and pluripotency of embryonic stem cells

Singh

Kagalwala

Parker-Thornburg³

et al. 2008

Nature

309

285

View full text Add to dashboard Cite

The neuronal repressor REST (RE1-silencing transcription factor; also called NRSF) is expressed at high levels in mouse embryonic stem (ES) cells 1 , but its role in these cells is unclear. Here we show that REST maintains self-renewal and pluripotency in mouse ES cells through suppression of the microRNA miR-21. We found that, as with known self-renewal markers, the level of REST expression is much higher in self-renewing mouse ES cells than in differentiating mouse ES (embryoid body, EB) cells. Heterozygous deletion of Rest (Rest 1/2 ) and its short-interfering-RNA-mediated knockdown in mouse ES cells cause a loss of self-renewal-even when these cells are grown under self-renewal conditions-and lead to the expression of markers specific for multiple lineages. Conversely, exogenously added REST maintains self-renewal in mouse EB cells. Furthermore, Rest 1/2 mouse ES cells cultured under self-renewal conditions express substantially reduced levels of several self-renewal regulators, including Oct4 (also called Pou5f1), Nanog, Sox2 and c-Myc, and exogenously added REST in mouse EB cells maintains the self-renewal phenotypes and expression of these self-renewal regulators. We also show that in mouse ES cells, REST is bound to the gene chromatin of a set of miRNAs that potentially target self-renewal genes. Whereas mouse ES cells and mouse EB cells containing exogenously added REST express lower levels of these miRNAs, EB cells, Rest 1/2 ES cells and ES cells treated with short interfering RNA targeting Rest express higher levels of these miRNAs. At least one of these REST-regulated miRNAs, miR-21, specifically suppresses the self-renewal of mouse ES cells, corresponding to the decreased expression of Oct4, Nanog, Sox2 and c-Myc. Thus, REST is a newly discovered element of the interconnected regulatory network that maintains the self-renewal and pluripotency of mouse ES cells.REST is believed to be a major transcriptional repressor of neurogenesis 2-5 , and activation of REST target genes was found to be sufficient to convert neural stem/progenitor cells to neuronal phenotypes 6,7 . However, REST activity seems to depend on the cellular context; for example, REST can show both an oncogenic 8-10 and tumour-suppressor function 5 as well as involvement in haematopoietic and cardiac differentiation [3][4][5] . Embryonic stem (ES) cells are pluripotent cells that have the potential for both indefinite selfrenewal and differentiation into all three germ layers of the body 11 . Here we provide evidence that REST has a unique role as a protector of self-renewal and pluripotency in mouse ES cells, corresponding to the expression of critical regulators such as Oct4, Nanog, Sox2 and c-Myc.We began by assessing the levels of REST protein in mouse ES cells growing under self-renewal conditions and differentiation conditions ( Fig. 1a; ES and EB, respectively). As expected, western blotting showed that the ES cells had higher levels of REST expression and of the representative markers of self-renewal (proteins Oct4, Sox2 and c-M...

show abstract

Section: Methods Summarymentioning

confidence: 99%

REST maintains self-renewal and pluripotency of embryonic stem cells

Singh

Kagalwala

Parker-Thornburg³

et al. 2008

Nature

309

285

View full text Add to dashboard Cite

show abstract

“…Eleven genes (OCT4, NANOG, SOX2, UTF1, DPPA5, LIN41, SOX1, H19, IGF2, GATA4, and HAND1) were selected to assess self-renewal and differentiation in human embryonic stem cell (Cai et al, 2006). OCT4, NANOG, UTF1, DPPA5, LIN41, SOX2 represent the undifferentiated hES cells' state.…”

Section: Real-time Qpcr Analysismentioning

confidence: 99%

Comparative evaluation of human embryonic stem cell lines derived from zygotes with normal and abnormal pronuclei

Qin¹,

Gao²,

Wang³

et al. 2010

Developmental Dynamics

View full text Add to dashboard Cite

Human embryonic stem (hES) cell lines have been derived from normally or abnormally fertilized zygotes. However, the similar and different properties of these two types of hES cell lines are not well-known. To address this question, we generated nine hES cell lines from zygotes containing normal (2PN) and abnormal (0PN, 1PN, 3PN) pronuclei. A side-by-side comparison showed that all cell lines exhibited distinct identity and karyotypical stability. They expressed similar ''stemness'' markers and alkaline phosphatase activity and differentiated into three embryonic germ lineages in embryoid bodies and teratomas. Under neural differentiation-promoting conditions, they were directed into neural progenitors and neurons. However, a variation in cell cycle and the relative abundance of gene expression of undifferentiated and differentiated markers were observed. These variations were also seen among individually derived normal hES cell lines. Thus, normal hES cell lines can be developed from fertilized zygotes with abnormal pronuclei usually excluded from clinical use. Developmental Dynamics 239:425-438,

show abstract

“…To avoid some of the cultivation dilemmas, regular karyotyping of the hESC line is required, as well as testing for possible mutations and the overall stability of the line, especially for long-term cultures with large number of passages. 68,69 Abnormalities in hESC karyotype have been reported in those cells that were cultured for long periods. 70 Ex vivo cultures with long duration of autologous stem cells may result in altered immunogenicity, which could lead to rejection of the cells upon transplantation.…”

Section: Limitationsmentioning

confidence: 99%

Therapeutic Potentials of Human Embryonic Stem Cells in Parkinson’s Disease

Newman

Bakay

2008

Neurotherapeutics

View full text Add to dashboard Cite

Summary:The loss of dopaminergic neurons of the substantia nigra is the pathological hallmark characteristic of Parkinson's disease (PD). The strategy of replacing these degenerating neurons with other cells that produce dopamine has been the main approach in the cell transplantation field for PD research. The isolation, differentiation, and long-term cultivation of human embryonic stem cells and the therapeutic research discovery made in relation to the beneficial properties of neurotrophic and neural growth factors has advanced the transplantation field beyond dopamine-producing cells. The present review addresses recent advances in human embryonic stem cell experimentation in relation to treating PD, as well as cell transplantation techniques in conjunction with alternative therapeutics.

show abstract

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

Cited by 127 publications

References 110 publications

REST maintains self-renewal and pluripotency of embryonic stem cells

REST maintains self-renewal and pluripotency of embryonic stem cells

Comparative evaluation of human embryonic stem cell lines derived from zygotes with normal and abnormal pronuclei

Therapeutic Potentials of Human Embryonic Stem Cells in Parkinson’s Disease

Contact Info

Product

Resources

About