• 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)

• BMB Reports
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• 제35권1호
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• pp.87-93
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• 2002

Neural cell survival is an essential concern in the aging brain and many diseases of the central nervous system. Neural transplantation of the stem cells are already applied to clinical trials for many degenerative neurological diseases, including Huntington's disease, Parkinson's disease, and strokes. A critical problem of the neural transplantation is how to reduce their apoptosis and improve cell survival. Neurotrophic factors generally contribute as extrinsic cues to promote cell survival of specific neurons in the developing mammalian brains, but the survival factor for neural stem cell is poorly defined. To understand the mechanism controlling stem cell death and improve cell survival of the transplanted stem cells, we investigated the effect of plausible neurotrophic factors on stem cell survival. The neural stem cell, HiB5, when treated with PDGF prior to transplantation, survived better than cells without PDGF. The resulting survival rate was two fold for four weeks and up to three fold for twelve weeks. When transplanted into dorsal hippocampus, they migrated along hippocampal alveus and integrated into pyramidal cell layers and dentate granule cell layers in an inside out sequence, which is perhaps the endogenous pathway that is similar to that in embryonic neurogenesis. Promotion of the long term-survival and differentiation of the transplanted neural precursors by PDGF may facilitate regeneration in the aging adult brain and probably in the injury sites of the brain.

• 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.

• 동의신경정신과학회지
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• 제20권4호
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• pp.63-77
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• 2009

Objectives : In this study, an essential oil fragrance from Danggwi was administrated into the neural stem cell and the effect of the essential oil on the differentiation and proliferation of the neural stem cells were observed. Methods : The establishment of the neural stem cell was identified via Nestin, DAPI dye. An essential oil fragrance from Danggwi was administrated with a proved optimum level for the survival of the cell through MTT assay. Also, according to the analysis of Western blot, the essential oil fragrance from Danggwi promotes the phosphorylating of Akt, Erk, ERM protein. Results : MTT assay showed increased in GFAP. The result indicates that the differentiation to astrocyte is promoted. The phosphorylation levels of ERM, Erk and Akt were increased at 60 min after addition of 5 ug/ml of essential oil fragrance from Danggwi and sustained to 48 hours. These imply that essential oil fragrance from Danggwi may induce the survival and the proliferation of the differentiated cells. Conclusions : These results suggest that the essential oil fragrance from Danggwi can be effective for the in vivo study of degenerative neuronal disease using neural stem cell.

• 대한생식의학회:학술대회논문집
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• 대한불임학회 2006년도 The 5th Biannual Meeting of Pacific Rim Society for Fertility and Sterility
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• pp.170.2-170.2
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• 2006

• Clinics in Shoulder and Elbow
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• 제15권2호
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• pp.65-72
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• 2012

목적: 세가지 기원의 줄기 세포 분화능과 면역표현형을 평가하고자 하였다. 대상 및 방법: 견봉하 점액낭과 골수, 탯줄 혈액 세 개의 군에서 세포를 채취하였다. 견봉하 점액낭과 골수는 견관절 수술 환자군에게 임상적 동의 하에 수술중 채취하였다. 각각의 채취된 세포 및 탯줄 혈액에 대하여 계대 배양을 시행하여 신경 분화군, 지방 분화군, 골 분화군을 평가하였으며 세포 표면 항체를 밝히기 위해 유동세포분석법을 이용하였다. 결과: 견봉하 점액낭 유래 세포에서는 신경분화와 지방 분화는 8예 모두 (100%)에서, 골분화는 8례 중 5예 (62.5%)에서 성공할 수 있었으며 골수 유래 세포의 경우 신경 및 지방 분화 유도한 6례 및 5예 모두 (100%) 분화에 성공하였으나 골분화 유도는 5예 중 4예 (80%)에서 얻을 수 있었다. 반면 탯줄 유래 세포 분화 연구의 경우 신경 분화 유도 67례 중 65예 (97%)에서 지방 분화 연구 54예 중 29예 (53.7%)에서 골 분화 연구 57예 중 39예 (68.4%)에서 성공할 수 있었다. 결론: 탯줄 유래 줄기세포의 분화능과 비교하였을 때 견봉하 점액낭 및 골수 유래 줄기세포의 분화능이 우수함을 알 수 있으며 이는 향후 세포 치료에 있어서 안정성 있는 치료 제공자가 될 수 있을 것으로 보이며 향후 생체 실험 연구의 참고 자료로서도 가치가 있을 것으로 보인다.

• 한국발생생물학회지:발생과생식
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• 제13권3호
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• pp.173-181
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• 2009

One of the most extensively studied populations of multipotent adult stem cells are mesenchymal stem cells (MSCs). MSCs derived from the human umbilical cord vein (HUC-MSCs) are morphologically and immunophenotypically similar to MSCs isolated from bone marrow. HUC-MSCs are multipotent stem cells, differ from hematopoietic stem cells and can be differentiated into neural cells. Since neural tissue has limited intrinsic capacity of repair after injury, the identification of alternate sources of neural stem cells has broad clinical potential. We isolated mesenchymal-like stem cells from the human umbilical cord vein, and studied transdifferentiation-promoting conditions in neural cells. Dopaminergic neuronal differentiation of HUC-MSCs was also studied. Neural differentiation was induced by adding bFGF, EGF, dimethyl sulfoxide (DMSO) and butylated hydroxyanisole (BHA) in N2 medium and N2 supplement. The immunoreactive cells for $\beta$-tubulin III, a neuron-specific marker, GFAP, an astrocyte marker, or Gal-C, an oligodendrocyte marker, were found. HUC-MSCs treated with bFGF, SHH and FGF8 were differentiated into dopaminergic neurons that were immunopositive for tyrosine hydroxylase (TH) antibody. HUC-MSCs treated with DMSO and BHA rapidly showed the morphology of multipolar neurons. Both immunocytochemistry and RT-PCR analysis indicated that the expression of a number of neural markers including NeuroD1, $\beta$-tubulin III, GFAP and nestin was markedly elevated during this acute differentiation. While the stem cell markers such as SCF, C-kit, and Stat-3 were not expressed after neural differentiation, we confirmed the differentiation of dopaminergic neurons by TH/$\beta$-tubulin III positive cells. In conclusion, HUC-MSCs can be differentiated into dopaminergic neurons and these findings suggest that HUC-MSCs are alternative cell source of therapeutic treatment for neurodegenerative diseases.

• Molecules and Cells
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• 제37권12호
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• pp.907-911
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• 2014

The function of reactive oxygen species (ROS) as second messengers in cell differentiation has been demonstrated only for a limited number of cell types. Here, we used a well-established protocol for BMP2-induced neuronal differentiation of neural crest stem cells (NCSCs) to examine the function of BMP2-induced ROS during the process. We first show that BMP2 indeed induces ROS generation in NCSCs and that blocking ROS generation by pretreatment of cells with diphenyleneiodonium (DPI) as NADPH oxidase (Nox) inhibitor inhibits neuronal differentiation. Among the ROS-generating Nox isozymes, only Nox4 was expressed at a detectable level in NCSCs. Nox4 appears to be critical for survival of NCSCs at least in vitro as down-regulation by RNA interference led to apoptotic response from NCSCs. Interestingly, development of neural crest-derived peripheral neural structures in Nox4-/- mouse appears to be grossly normal, although Nox4-/- embryos were born at a sub-Mendelian ratio and showed delayed over-all development. Specifically, cranial and dorsal root ganglia, derived from NCSCs, were clearly present in Nox4-/- embryo at embryonic days (E) 9.5 and 10.5. These results suggest that Nox4-mediated ROS generation likely plays important role in fate determination and differentiation of NCSCs, but other Nox isozymes play redundant function during embryogenesis.

• Clinical and Experimental Reproductive Medicine
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• 제29권2호
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• pp.117-127
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• 2002

Objective: This study was to investigate the generation of the functional neuron derived from human embryonic stem (hES, MB03) cells on in vitro neural cell differentiation system. Methods: For neural progenitor cell formation derived from hES cells, we produced embryoid bodies (EB: for 5 days, without mitogen) from hES cells and then neurospheres (for $7{\sim}10$ days, 20 ng/ml of bFGF added N2 medium) from EB. And then finally for the differentiation into mature neuron, neural progenitor cells were cultured in i) N2 medium only (without bFGF), ii) N2 supplemented with 20 ng/ml platelet derived growth factor-bb (PDGF-bb) or iii) N2 supplemented with 5 ng/ml brain derived neurotrophic factor (BDNF) for 2 weeks. Identification of neural cell differentiation was carried out by immunocytochemistry using $\beta_{III}$-tubulin (1:250), MAP-2 (1:100) and GFAP (1:500). Also, generation of functional neuron was identified using anti-glutamate (Sigma, 1:1000), anti-GABA (Sigma, 1:1000), anti-serotonin (Sigma, 1:1000) and anti-tyrosine hydroxylase (Sigma, 1:1000). Results: In vitro neural cell differentiation, neurotrophic factors (PDGF and BDNF) treated cell groups were high expressed MAP-2 and GFAP than non-treated cell group. The highest expression pattern of MAP-2 and $\beta_{III}$-tubulin was indicated in BDNF treated group. Also, in the presence of PDGF-bb or BDNF, most of the neural cells derived from hES cells were differentiated into glutamate and GABA neuron in vitro. Furthermore, we confirmed that there were a few serotonin and tyrosine hydroxylase positive neuron in the same culture environment. Conclusion: This results suggested that the generation of functional neuron derived from hES cells was increased by addition of neurotrophic factors such as PDGF-bb or BDNF in b-FGF induced neural cell differentiation system and especially glutamate and GABA neurons were mainly produced in the system.

• Reproductive and Developmental Biology
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• 제28권4호
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• pp.247-252
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• 2004

Human embryonic stem (ES) cells are derived from the inner cell mass of the preimplantation embryo. Human ES cells have the capacity to differentiate into various types of cells in the body. Human ES cells are indefinite source of cells for cell therapy in various degenerative disorders including neuronal disorders. Directed differentiation of human ES cells is a prerequisite for their clinical application. The objective of this study is to develop the culture condition for the derivation of neural precursor cells from human ES cells. Neural precursor cells were derived from human ES cells in a stepwise culture condition. Neural precursor cells in the form of neural rosette structures developed into neurospheres when cultured in suspension. Suspension culture of neurospheres has been maintained over 4 months. Expressions of nestin, soxl, sox2, pax3 and pax6 transcripts were upregulated during differentiation into neural precursor cells by RT-PCR analysis. In contrast, expression of oct4 was dramatically downregulated in neural precursor cells. Immunocytochemical analyses of neural precursor cells demonstrated expression of nestin and SOX1. When induced to differentiate on an adhesive substrate, neuro-spheres were able to differentiate into three lineages of neural systems, including neurons, astrocytes and oligo-dendrocytes. Transcripts of sox1 and pax6 were downregulated during differentiation of neural precursor cells into neurons. In contrast, expression of map2ab was elevated in the differentiated cells, relative to those in neural precursor cells. Neurons derived from neural precursor cells expressed NCAM, Tuj1, MAP2ab, NeuN and NF200 in immunocytochemical analyses. Presence of astrocytes was confirmed by expression of GFAP immuno-cytochemically. Oligodendrocytes were also observed by positive immuno-reactivities against oligodendrocyte marker O1. Results of this study demonstrate that a stepwise culture condition is developed for the derivation of neural precursor cells from human ES cells.