• 한국미생물·생명공학회지
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• 제39권4호
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• pp.364-373
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• 2011

인간배아줄기세포는 인간배아줄기세포가 가지는 전 분화능 등의 특이적 특성으로 인해 재생의학 분야에서 세포 치료제의 근원으로 널리 각광받고 있다. 그러나, 미분화 상태의 인간배아줄기세포를 세포치료제로 이용하기 위해서는 인간배아줄기세포 주 유래 기능성 세포를 확립이 반드시 요구된다. 본 연구에서는, 미분화 상태의 인간배아줄기세포주로부터 기능성 세포의 확립을 위해, 혈관계통의 세포로 분화를 유도하였으며, 분화 유도 후 세포의 크기 차이를 이용하여 특정 세포군 만을 분리하여 그 기능성을 비교 분석하였다. 그 결과, VEGF를 이용하여 분화 시킨 세포군에서 약 10%의 PECAM 양성 세포군을 확인할 수 있었으며, 분리 및 세포 이식을 위해 세포를 단일 세포군으로 만들었다. 단일 세포군의 형성 후, 유세포 분석기를 이용한 세포 분리 기법을 이용하여 FCS를 기준으로 한 세포 크기의 차이를 이용하여 특정 세포군 만을 분리하여, 하지 허혈 동물 모델로의 이식을 통해, 비 분리 세포군과 치료 효능을 비교 분석을 실시하였다. 세포 이식 4주 후, 혈류량 복구율이 FSC 기준 분리 군의 경우 54%, 비 분리군의 경우 17%를 보이는 것을 확인하였다. 이 결과는, 초기 분화 유도 후 세포 크기차이를 이용한 세포 분리법이 기능성 세포 획득에 이용될 수 있음을 시사한다. 이와 같은 방법을 통해 다양한 종류의 기능성 세포 분리에 이용될 수 있을 것이라 생각된다.

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2002년도 춘계학술발표대회 발표논문초록집
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• pp.19-19
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• 2002

This study was to examine whether the in vitro differentiated neural cells derived from human embryonic stem (hES, MB03) cells can be survived and expressed tyrosin hydroxylase(TH) in grafted normal or PD rat brain. To differentiate in vitro into neural cells, embryoid bodies (EB: for 5 days, without mitogen) were formed from hES cells, neural progenitor cells(neurosphere, for 7-10 days, 20 ng/㎖ of bFGF added N2 medium) were produced from EB, and then finally neurospheres were differentiated into mature neuron cells in N2 medium(without bFGF) for 2 weeks. In normal rat brain, neural progenitor cells or mature neuron cells (1×10/sup 7/ cells/㎖) were grafted to the striatum of normal rats. After 2 weeks, when the survival of grafted hES cells was examined by immunohistochemical analysis, the neural progenitor cell group indicated higher BrdU, NeuN＋, MAP2＋ and GFAP＋ than mature neuron cell group in grafted sites of normal rats. This result demonstrated that the in vivo differentiation of grafted hES cells be increased simultaneously in both of neuronal and glial cell type. Also, neural progenitor cell grafted normal rats expressed more TH pattern than mature neuron cells. Based on this data, as a preliminary test, when the neural progenitor cells were grafted into the striatum of 6-hydroxydopamine lesioned PD rats, we confirmed the cell survival (by double staining of Nissl and NeuN) and TH expression. This result suggested that in vitro differentiated neural progenitor cells derived from hES cells are more usable than mature neuron cells for the neural cell grafting in animal model and those grafted cells were survived and expressed TH in normal or PD rat brain.

• Reproductive and Developmental Biology
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• 제29권2호
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• pp.93-99
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• 2005

Human embryonic stem cells (hESCs) derived from the inner cell mass of blastocysts have the ability to renew themselves and to differentiate into cell types of all lineage. The present study was carried out to investigate whether the Wnt signaling pathway is related to maintaining self-renewal of hESCs. Glycogen Synthase Kinase 3 (GSK-3) inhibitor, BIO ((2'Z,3'E)-6-Bromoindirubin-3'-oxime) was treated to Miz-hES1 line for activation of Wnt signaling pathway. BIO-nontreated hESCs (control) and BID-treated hESCs were cultured for 5 days in the modified feeder-free system. During the culture of hESCs, differences were observed in the colony morphology between 2 groups. Controls were spread outwards whereas BIO-nontreated hESCs were clumped in the center and the differentiated cells were spreading outwards in the edges. The results of stem cell specific marker staining indicated that control were differentiated in large part whereas BIO-treated hESCs maintain self-renewal in the center of the colony. The results of lineage marker staining suggested that outer cells of the hESC colony were differentiated to the neuronal progenitor cells in both control and BIO-treated hESC. These results indicate that Wnt signaling is related to self-renewal in hESCs. In addition, control group showed higher composition of apoptotic cells $(23.76\%)$ than the BID-treated group $(5.59\%)$. These results indicate that BIO is effective on antapoptosis of hESCs.

• 생체재료학회지
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• 제22권4호
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• pp.311-318
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• 2018

Background: Tissue regeneration includes delivering specific types of cells or cell products to injured tissues or organs for restoration of tissue and organ function. Stem cell therapy has drawn considerable attention since transplantation of stem cells can overcome the limitations of autologous transplantation of patient's tissues; however, it is not perfect for treating diseases. To overcome the hurdles associated with stem cell therapy, tissue engineering techniques have been developed. Development of stem cell technology in combination with tissue engineering has opened new ways of producing engineered tissue substitutes. Several studies have shown that this combination of tissue engineering and stem cell technologies enhances cell viability, differentiation, and therapeutic efficacy of transplanted stem cells. Main body: Stem cells that can be used for tissue regeneration include mesenchymal stem cells, embryonic stem cells, and induced pluripotent stem cells. Transplantation of stem cells alone into injured tissues exhibited low therapeutic efficacy due to poor viability and diminished regenerative activity of transplanted cells. In this review, we will discuss the progress of biomedical engineering, including scaffolds, biomaterials, and tissue engineering techniques to overcome the low therapeutic efficacy of stem cells and to treat human diseases. Conclusion: The combination of stem cell and tissue engineering techniques overcomes the limitations of stem cells in therapy of human diseases, and presents a new path toward regeneration of injured tissues.

• 한국발생생물학회:학술대회논문집
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• 한국발생생물학회 2004년도 추계학술대회
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• pp.22-22
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• 2004

• Reproductive and Developmental Biology
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• 제33권3호
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• pp.133-137
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• 2009

Pluripotent embryonic stem (ES) cells isolated from inner cell mass (ICM) of blastocyst-stage embryos are capable of differentiating into various cell lineages and demonstrate germ-line transmission in experimentally produced chimeras. These cells have a great potential as tools for transgenic animal production, screening of newly-developed drugs, and cell therapy. Miniature pigs, selectively bred pigs for small size, offer several advantages over large breed pigs in biomedical research including human disease model and xenotransplantation. In the present study, factors affecting primary culture of somatic cell nuclear transfer blastocysts from miniature pigs for isolation of ES cells were investigated. Formation of primary colonies occurred only on STO cells in human ES medium. In contrast, no ICM outgrowth was observed on mouse embryonic fibroblasts (MEF) in porcine ES medium. Plating intact blastocysts and isolated ICM resulted in comparable attachment on feeder layer and primary colony formation. After subculture of ES-like colonies, two putative ES cell lines were isolated. Colonies of putative ES cells morphologically resembled murine ES cells. These cells were maintained in culture up to three passages, but lost by spontaneous differentiation. The present study demonstrates factors involved in the early stage of nuclear transfer ES cell isolation in miniature pigs. However, long-term maintenance and characterization of nuclear transfer ES cells in miniature pigs are remained to be done in further studies.

• 한국발생생물학회:학술대회논문집
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• 한국발생생물학회 2004년도 추계학술대회
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• pp.21-21
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• 2004

• 한국수정란이식학회:학술대회논문집
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• 한국수정란이식학회 2004년도 제4회 발생공학 국제심포지움 및 학술대회
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• pp.88-88
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• 2004

• 한국수정란이식학회:학술대회논문집
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• 한국수정란이식학회 2004년도 제4회 발생공학 국제심포지움 및 학술대회
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• pp.98-99
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• 2004

• Reproductive and Developmental Biology
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• 제35권1호
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• pp.1-8
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• 2011

Techniques to evaluate gene expression profiling, such as sufficiently sensitive cDNA microarrays or real-time quantitative PCR, are efficient methods for monitoring human pluripotent stem cell (hESC/iPSC) cultures. However, most of these high-throughput tests have a limited use due to high cost, extended turn-around time, and the involvement of highly specialized technical expertise. Hence, there is an urgency of rapid, cost-effective, robust, yet sensitive method development for routine screening of hESCs/hiPSCs. A critical requirement in hESC/hiPSC cultures is to maintain a uniform undifferentiated state and to determine their differentiation capacity by showing the expression of gene markers representing all three germ layers, including ectoderm, mesoderm, and endoderm. To quantify the modulation of gene expression in hESCs/hiPSC during their propagation, expansion, and differentiation via embryoid body (EB) formation, we developed a simple, rapid, inexpensive, and definitive multimarker, semiquantitative multiplex RT-PCR platform technology. Among the 9 gene primers tested, 5 were pluripotent markers comprising set 1, and 3 lineage-specific markers were combined as set 2, respectively. We found that these 2 sets were not only effective in determining the relative differentiation in hESCs/hiPSCs, but were easily reproducible. In this study, we used the hES/hiPS cell lines to standardize the technique. This multiplex RT-PCR assay is flexible and, by selecting appropriate reporter genes, can be designed for characterization of different hESC/hiPSC lines during routine maintenance and directed differentiation.