• 한국발생생물학회지:발생과생식
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• 제15권1호
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• pp.41-52
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• 2011

In the present study, embryoid bodies (EBs) obtained from induced pluripotent stem cells (iPSCs) were induced to differentiate into germ lineage cells by treatment with bone morphogenetic protein 4 (BMP4) and retinoic acid (RA). The results were compared to the results for embryonic stem cells (ESCs) and multipotent spermatogonial stem cells (mSSCs) and quantified using immunocytochemical analysis of germ cell-specific markers (integrin-${\alpha}6$, GFR-${\alpha}1$, CD90/Thy1), fluorescence activating cell sorting (FACS), and real time-RT-PCR. We show that the highest levels of germ cell marker-expressing cells were obtained from groups treated with 10 ng/$m{\ell}$ BMP4 or 0.01 ${\mu}M$ RA. In the BMP4-treated group, GFR-${\alpha}1$ and CD90/Thy-1 were highly expressed in the EBs of iPSCs and ESCs compared to EBs of mSSCs. The expression of Nanog was much lower in iPSCs compared to ESCs and mSSCs. In the RA treated group, the level of GFR-${\alpha}1$ and CD90/Thy-1 expression in the EBs of mSSCs Induced pluripotent stem cells, Mouse embryonic stem cells, Multipotent spermatogonial stem cells, Germ cell lineage, Differentiation potential. was much higher than the levels found in the EBs of iPSCs and similar to the levels found in the EBs of ESCs. FACS analysis using integrin-${\alpha}6$, GFR-${\alpha}1$, CD90/Thy1 and immunocytochemistry using GFR-${\alpha}1$ antibody showed similar gene expression results. Therefore our results show that iPSC has the potential to differentiate into germ cells and suggest that a protocol optimizing germ cell induction from iPSC should be developed because of their potential usefulness in clinical applications requiring patient-specific cells.

• Animal cells and systems
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• 제13권2호
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• pp.235-245
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• 2009

Embryonic stem (ES) cells have a capability to generate all types of cells. However, the mechanism by which ES cells differentiate into specific cell is still unclear. Using microarray technology, the differentiation process in mouse embryonic stem cells was characterized by temporal gene expression changes of mouse ES cells during differentiation in a monolayer culture. A large number of genes were differentially regulated from 1 day to 14 days, and less number of genes were differentially expressed from 14 days to 28 days. The number of up-regulated genes was linearly increased throughout the 28 days of in vitro differentiation, while the number of down-regulated genes reached the plateau from 14 days to 28 days. Most differentially expressed genes were functionally classified into transcriptional regulation, development, extra cellular matrix (ECM),cytoskeleton organization, cytokines, receptors, RNA processing, DNA replication, chromatin assembly, proliferation and apoptosis related genes. While genes encoding ECM proteins were up-regulated, most of the genes related to proliferation, chromatin assembly, DNA replication, RNA processing, and cytoskeleton organization were down-regulated at 14 days. Genes known to be associated with embryo development or transcriptional regulation were differentially expressed mostly after 14 days of differentiation. These results indicate that the altered expression of ECM genes constitute an early event during the spontaneous differentiation, followed by the inhibition of proliferation and lineage specification. Our study might identify useful time-points for applying selective treatments for directed differentiation of mouse ES cells.

• BMB Reports
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• 제41권10호
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• pp.705-709
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• 2008

NF-${\kappa}B$ is a transcriptional regulator involved in many biological processes including proliferation, survival, and differentiation. Recently, we reported that expression and activity of NF-${\kappa}B$ is comparatively low in undifferentiated human embryonic stem (ES) cells, but increases during differentiation. Here, we found a lower expression of NF-${\kappa}B$ p65 protein in mouse ES cells when compared with mouse embryonic fibroblast cells. Protein levels of NF-${\kappa}B$ p65 and relB were clearly enhanced during retinoic acid-induced differentiation. Furthermore, increased DNA binding activity of NF-${\kappa}B$ in response to TNF-$\alpha$, an agonist of NF-${\kappa}B$ signaling, was seen in differentiated but not undifferentiated mouse ES cells. Taken together with our previous data in human ES cells, it is likely that NF-${\kappa}B$ expression and activity of the NF-${\kappa}B$ signaling pathway is comparatively low in undifferentiated ES cells, but increases during differentiation of ES cells in general.

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2004년도 춘계학술발표대회
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• pp.276-276
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• 2004

Pluripotent stem cells have been generated from two embryonic sources. ES cells are generated from ICM of blastocyst stage embryos, and embryonic germ (EG) cells are generated from primordial germ cells (PGCs). Both ES and EG cells are pluripotent and present important characteristics such as high levels of alkaline phosphatase (AP) activity, multi-cellular colony formation, normal and stable karyotypes, continuously passaging ability, and the capability of differentiation into all three embryonic germ layers. (omitted)

• Reproductive and Developmental Biology
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• 제36권1호
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• pp.21-26
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• 2012

Suspension culture is a useful tool for culturing embryonic stem (ES) cells in large-scale, but the stability of pluripotency and karyotype has to be maintained $in$ $vitro$ for clinical application. Therefore, we investigated whether the chromosomal abnormality of ES cells was induced in suspension culture or not. The ES cells were cultured in suspension as a form of aggregate with or without mouse embryonic fibroblasts (MEFs), and 0 or 1,000 U/ml leukemia inhibitory factor (LIF) was treated to suspended ES cells. After culturing ES cells in suspension, their karyotype, DNA content, and properties of pluripotency and differentiation were evaluated. As a result, the formation of tetraploid ES cell population was significantly increased in suspension culture in which ES cells were co-cultured with both MEFs and LIF. Tetraploid ES cell population was also generated when ES cells were cultured alone in suspension regardless of the existence of LIF. On the other hand, the formation of tetraploid ES cell population was not detected in LIF-free condition, in which MEFs were included. The origin of tetraploid ES cell population was turned out to be E14 ES cells and not MEFs by microsatellite analysis and the basic properties of them were still maintained despite ploidy-conversion to tetraploidy. Furthermore, we identified the ploidy shift from tetraploidy to near-triploidy as tetraploid ES cells were differentiated spontaneously. From these results, we demonstrated that suspension culture system could induce ploidy-conversion generating tetraploid ES cell population. Moreover, optimization of suspension culture system may make possible mass-production of ES cells.

• 한국발생생물학회:학술대회논문집
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• 한국발생생물학회 2007년도 전기 24차 학술대회
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• pp.3-3
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• 2007

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2004년도 춘계학술발표대회
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• pp.273-273
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• 2004

This study was to examine in vitro neural cell differentiation pattern of the genetically modified human embryonic stem cells expressing tyrosine hydroxylase (TH). Human embryonic stem (hES, MB03) cell was transfected with cDNAs cording for TH. Successful transfection was confirmed by western immunoblotting. (omitted)

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2003년도 학술발표대회 발표논문초록집
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• pp.74-74
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• 2003

Embryonic stem cells have several characteristics suitable for cell replacement therapy. To investigate a possibility of using human embryonic stem cell (hESC) as a carrier of therapeutic gene(s), hESC (MB03) was co-transfected with cDNAS coding for tyrosine hydroxylase (TH) and GTP cyclohydrolase Ⅰ (GTPCH Ⅰ) and bulk-selected using neomycin and hygromycin-B. Successful transfection was confirmed by western immunoblotting and RT-PCR. The genetically modified hESC (bk-THGC) relieved apomorphine-induced asymmetric motor behavior by approximately 54% when grafted into striatum of 6-OHDA-denervated rat brain. The number of rotation, however, increased up to 176+18% in 6 weeks when sham-grafted compared with number of rotation before graft. Immunohistochemical staining revealed that the grafted hESC survived and expressed TH for at least 6 weeks while the experiment was continued.

• Molecules and Cells
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• 제42권3호
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• pp.200-209
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• 2019

Embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) have been used as promising tools for regenerative medicine, disease modeling, and drug screening. Traditional and common strategies for pluripotent stem cell (PSC) differentiation toward disease-relevant cell types depend on sequential treatment of signaling molecules identified based on knowledge of developmental biology. However, these strategies suffer from low purity, inefficiency, and time-consuming culture conditions. A growing body of recent research has shown efficient cell fate reprogramming by forced expression of single or multiple transcription factors. Here, we review transcription factor-directed differentiation methods of PSCs toward neural, muscle, liver, and pancreatic endocrine cells. Potential applications and limitations are also discussed in order to establish future directions of this technique for therapeutic purposes.

• 한국가금학회지
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• 제27권1호
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• pp.73-78
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• 2000

Embryonic stem (ES) cells are pluripotent cell lines, which derived from preimplantation embryo. These cells have been used as a vehicle of foreign DNA for production of transgenic mammals. this experiment was performed to examined the possible use of blastodermal cells derived from hen's egg for germline manipulation. Stage X blsdtodermal cells isolated from fertilized eggs were cultured in DMEM containing 15% fetal calf serum. Blastodermal cells wre co-cultured on the chicken embryonic fibroblast (CEF) or mouse embryonic fibroblast(MEF) cells. to examine the effects of growth factors on stem cell growth, bFGF and LIF were added. There was no significant difference in colony formation of putative ES cells between CEF and MEF as a feederlayer, but the addition of growth factors enhanced the proliferation and inhibited differentiation of blastodermal cells. To characterize the cell colonies as a putative ES cells, putative embryonic cell colonies were stained by periodic acid Schiffs (PAS) reagent. The putative ES cell colonies showed intensive positive reaction similar to the property of undifferentiated PGC upto 20days in vitro, but not in other cell types. this result demonstrates that PAS-positive cell colonies may be used for the study of establishment of chicken ES cell lines for the production of transgenic chicken.