• Proceedings of the KSAR Conference
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• 2004.06a
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• pp.271-271
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• 2004

본 실험은 TGF-a를 처리하여 분화가 유도된 인간배아 줄기세포를 파킨슨 동물모델에 이식하여 숙주세포에서의 생존 및 이식효과를 검토하고자 실시하였다. TGF-a로 분화된 세포의 이식효과를 판정하고자 배양시 TGF-a처리군과 처리하지 않은 군으로 나누어 분화를 유도한 인간배아 줄기세포를 hoechst33342로 표지 하여 병변 유발과 동일한 방법으로 동측 선조체내에 4×10⁴개/2ul가 되도록 이식하고(이식 위치: AP 0.7, ML 2.0, DV3.4) 이식 후 2, 4주에서 행동학적 변화를 관찰하고 4주에 동물을 희생시켜 4% PFA를 이용하여 뇌 조직을 고정하고 뇌 조직은 40㎛ 두께로 동결 절편을 만들어 면역조직화학염색을 시행하여 신경세포로의 분화 및 TH 발현 여부를 관찰하였고 분화의 표지물질로 nestin, NF200, GFAP, TH를 사용하여 형태학적 변화를 관찰하였다. (중략)

• Journal of Science and Technology Studies
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• v.12 no.1
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• pp.185-207
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• 2012

Since the Hwang scandal, the South Korean state has expressed often-conflicting interests of encouraging stem cell research and the IVF industry to save the country and introducing the ethical regulation in conformity with "Global Standard." As the tightening ethical regulation of stem cell research has enervated the field of human Embryonic stem cell(hESC) research, somatic stem cells (re-)emerged as an alternative savior that could rescue the future of research communities, bio-industry, practicing doctors, patients and the nation itself from the crisis. The recent literature on Korean biotechnology, however, mainly focus on hESC and relatively little attention has been given to the rapidly growing field of research on somatic stem cells like hematopoietic stem cells(HSCs) or Adipose derived stem cells(ASCs). While the hESC therapy is often regarded as experimental and ethically controversial, the HSCs or Mesenchymal stem cell(MSC) therapies have already made their ways into people's everyday life through market without much public discussion. Many ordinary people in South Korea are familiar with the story of patients who survived leukemia with the HSCs treatment; the number of doctors who are actively marketing the ASCs therapies is on the rapid increase; the concept of cosmetic products made from ASCs is gaining popularity among consumers. In this context, this article argues that the current ethical debates solely focusing on hESC or on the state policy and research regulation are too limiting to fully illuminate the politics of stem cell technologies in South Korea.

• Journal of Convergence for Information Technology
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• v.8 no.2
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• pp.61-65
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• 2018

Our experiment studied that grafted stem cells reduced behavioral deficiency in rodent animal models of clip compressive surgery inducing spinal cord infarction. Our research proved the effect of embryonic stem cells to the spinal cord infarction caused by compressing T9-10 with an aneurysm clip, focusing the application of grafted stem cells for reduction of infarction and regeneration of spinal cord nervous injury. Therefore, our research suggests manifest results that implantation of mouse embryonic stem cell could show behavioral improvement after severe spinal cord damage. Therefore, mouse embryonic stem cell (mESC) could be useful application for the method in neurological injury. Conclusively, stem cell implant therapy may enhance the effectiveness of stem cell implant for central nervous system injury.

• Clinical and Experimental Reproductive Medicine
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• v.36 no.4
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• pp.283-292
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• 2009

Objective: Human embryonic stem cells (hESCs) can proliferate indefinitely and differentiate into all kinds of cell types in vitro. Therefore, hESCs can be used as a cell source for cell-based therapy. Transduction of foreign genes to hESCs could be useful for tracing differentiation processes of hESCs and elucidation of gene function. Thus, we tried to introduce enhanced green fluorescent protein (eGFP) gene to hESCs, XX and XY cell lines in this study. Methods: Lentivirus containing eGFP was packaged in 293T cells and applied to hESCs to transduce eGFP. Expression of transduced eGFP was evaluated under the fluorescence microscope and eGFP positive population was analyzed by FACS. Expression of undifferentiation state markers such as Oct4, Nanog, SSEA4 and Tra-1-81 was examined by RT-PCR and/or immunofluorescence in eGFP-hESCs after transduction. In addition, the ability of eGFP-hESCs to form embryoid bodies (EBs) was tested. Results: eGFP was successfully transduced to hESCs by lentivirus. eGFP expression was stably maintained up to more than 40 passages. eGFP-hESCs retained expression patterns of undifferentiation state markers after transduction. Interestingly, disappearance of transduced eGFP was notably observed during spontaneous differentiation of eGFP-hESCs. Conclusion: We established eGFP expressing hESC lines using lentivirus and showed the maintenance of undifferentiation characteristics of these eGFP-hESCs. This reporter-containing hESCs could be useful for tracing the processes of differentiation of hESCs and other studies.

• Clinical and Experimental Reproductive Medicine
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• v.35 no.1
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• pp.39-48
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• 2008

Objective: The aim of this study was to investigate whether multipotent germline stem cells (MGSCs) can be established from neonatal mouse testis. Methods: Various cells containing MGSCs were collected from neonatal testis of ICR mice and allocated to plates for in vitro culture. After 7 days in culture, the cells were passed to a fresh culture plate and continuously cultured. From the third or fourth passage, the presumed MGSCs were cultured and maintained on mitomycin C-inactivated STO feeder cells. The MGSCs were cultured in a condition where mouse embryonic stem cells (ESCs) are cultured. Characteristics of the MGSCs were evaluated by RT-PCR, immunocytochemistry, alkaline phosphatase activity, karyotyping, and transmission electron microscopy. Results: Two MGSCs lines were established from 9 pooled sets of neonatal testicular cells. MGSCs colonies were morphologically undistinguishable from ESCs colonies and both MGSC lines as well as ESCs expressed undifferentiated stem cell markers, such as Thy-1, Oct-4, Nanog, Sox2 and alkaline phosphatase. Fine structure of undifferentiated MGSCs were similar to those of ESCs and 60% of MGSCs (12/20) had normal karyotype at passage 10. They were able to form embryoid bodies (EBs) and MGSC-derived EBs expressed marker genes of three germ layers. Conclusion: We could establish the MGSCs from neonatal mouse testis and they were differentiated to multipotent lineages of three germ layers. Molecular characteristics of MGSCs were similar to those of ESCs. Our results suggest a possibility that multipotent stem cells derived from testis, the MGSCs, could replace the ESCs in biotechnology and regenerative medicine.

• Development and Reproduction
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• v.11 no.2
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• pp.111-119
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• 2007

Differentiation of cells can be induced through modulation of endogenous regulators using exogenous factors. Useful transfection systems to transport a specific exogenous regulator into cell have been tried but still there are many obstacles to overcome. In this study, we examined the transfection efficiency of cell permeable peptides (CPPs) in mouse embryonic stem cell under the various conditions. To identify the CPP-mediated translocation of a protein, we employed recombinant CPP-enhanced green fluorescent protein (EGFP). Viability of R1 cells was different between experimental groups depending on the kind of CPP and the concentration of CPP-EGFP. Translocation of CPP-EGFPs into the R1 cells was not detected until 30 min after CPP-EGFPs treatment in all groups. After 1 hr, translocation of pEP-1-EGFP-N was detected, but it could not be detected in the other group. Transfection of pEP-1EGFP-N was independent on its concentration. The time course did not show saturation even after 24 hr in pEP-1-EGFP-N. These results showed that the permeability depended on the kind of CPP and the location of His-tag in the case of examined CPPs, and did not need biological energy. On summary, the efficiency of transfection of CPP-EGFP depends on the CPP sequences but the culture time is not a key factor in transfection for the mouse embryonic stem cell. For the future studies to improve the efficiency of translocation of protein into embryonic stem cells, it is needed to develop modified CPP or mediator. The studies would be very useful to induce the differentiation of embryonic stem cells.

• Development and Reproduction
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• v.14 no.2
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• pp.43-50
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• 2010

Recently induced pluripotent stem (iPS) cells are derived from somatic cells by ectopic expression of several transcription factors (reprogramming factors) using technology of somatic cell reprogramming. iPS cells are able to selfrenew and differentiate into all type of cells in the body similarly to embryonic stem cells. Because iPS cells have advantages that can avoid immune rejection after transplantation and ethical issues unlike embryonic stem cells, research on iPS has made significant progress since the first report by Yamanaka in 2006. Nevertheless of many advantages of iPS, safer methods to introduce reprogramming factors into somatic cells must be developed due to safety concerns regarding viral vectors, and safer reprogramming factors to substitute the oncogenes should be evaluated for clinical application of iPS. Here we discuss the recent progress in reprogramming factors in embryonic stem cells, oocytes, and embryos, and discuss further research for finding new, more reliable and safer reprogramming factors.

• Clinical and Experimental Reproductive Medicine
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• v.31 no.1
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• pp.41-50
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• 2004

연구 목적: 본 연구는 인간배아줄기세포 동결에 minimum volume cooling (MVC) 초자화 동결방법이 유용하게 이용될 수 있는지의 여부를 조사하고자 실시하였다. 연구재료 및 방법: 인간배아줄기세포 콜로니는 0.05% collagenase 처리와 기계적 처리에 의해 작은 clumps로 자른 다음 동결 방법에 따른 효율을 비교 검토하고자, i) 대략 40-50개의 clumps를 10% DMSO가 들어있는 동결액에 $5{\sim}10$분간 처리하여 1ml cryo-vial에 넣고 slow-cooling용 cryo-module에 장착하고 -80C에서 overnight한 후 $LN_2$에 침지하여 완만동결을 실시하거나 ii) 10% ethylene glycol (EG)이 들어 있는 동결액에서 5$\sim$10분 처리하고 30% EG과 0.5 mol sucrose가 들어 있는 동결액에서 30초간 처리하여 본 연구를 위해 개발된 MVC straw에 10 clumps씩 적재한 다음 $4{\sim}5$ MVC straw를 $LN_2$가 들어있는 cryo-vial에 넣어 MVC 초자화동결을 실시하였다. 융해 후 생존율을 조사하였고 배아줄기세포의 특성을 유지하고 있는지의 여부를 조사하였다. 결 과: 융해 후, 인간배아줄기세포의 생존율은 완만동결을 실시했던 군 (20.0%) 보다 MVC 초자화동결을 실시했던 군에서 (76.0%) 유의하게 높게 나타났다. 또한 회복과정에서 완만동결을 실시했던 군에서는 세포성장이 매우 더디고 안 좋은 반면, MVC 초자화동결을 실시했던 군은 배아줄기세포의 증식이 동결을 실시하지 않은 세포와 같은 상태로 2주 이후부터 빠르게 전환되고 회복되는 것을 확인할 수 있었다. 이와 더불어 MVC 초자화동결－융해 후 회복된 배아줄기세포에서 정상 핵형, alkaline phosphatase acitivity, SSEA-4와 TRA-1-60 염색 및 Oct-4 발현을 확인하였으며 체외분화의 특성도 확인하였다. 결 론: 새로이 개발된 MVC 초자화동결을 이용하면 인간배아줄기세포는 고유의 특성을 잃지 않고 성공적으로 동결될 수 있다.

• The Journal of the Korean life insurance medical association
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• v.25
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• pp.49-62
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• 2006

생명공학의 시대로 일컬어 지고 있는 오늘날, 재생의학 분야에서는 난치성 질환 치료를 목적으로 활발한 연구가 진행되고 있다. 특히, 줄기세포를 이용한 세포 대체치료 관련 연구는 최근 국내 황우석 박사의 체세포 핵이식 배아세포주 확립에 이르기 까지 괄목할 만한 발전을 보여 주고 있다. 이와 관련해 생명보험산업에 적잖은 파장이 예상되며, 생명보험사 내부적으로 기존에 판매된 상품의 사차손 관리와 함께 급속도로 발전하는 줄기세포 연구에 직접적으로 대응하는 상품개발, 언더라이팅, 지급 심사 등 보험사 내외에서의 전방위적인 변화가 필요하다는 문제 제기가 있다. 줄기세포란 조직 분화 과정에서 볼 수 있는 세포이며 근육 뼈 뇌 피부 등 신체의 어떤 기관으로도 전환할 수 있는 만능세포로서, 간 폐 심장 등 구체적 장기를 형성하기 이전에 분화를 멈출 배아 단계의 세포를 말한다. 한편, 성체줄기세포는 조직이나 기관의 분화된 세포들 사이에서 발견되는 미분화 세포로써, 자기 스스로 증식할 수 있으며, 조직이나 기관의 특수한 기능을 가지고 있는 세포로 분화할 수 있는 능력을 가진 신체줄기세포를 말한다. 배아줄기세포와 생체줄기세포를 통한 장기이식 등 난치병 정복은 윤리적, 사회적으로 많은 논란이 예상되며, 기술적으로도 해결해야 할 문제점들이 산적해 있기 때문에 아직은 요원한 것이 사실이다. 현재 유럽 대부분의 나라와 미국에서는 인간 배아의 복제가 금지되어 있으며, 일본 정부는 연구용 배아 복제를 제한적으로 허용하고 있다. 하지만, 우리 나라의 경우 2005년 1월에 '생명윤리 및 안전에 관한 법'이 발효되었지만 정부는 관련 부작용에 대한 깊은 고찰 없이 전폭적인 지원들 약속하고 있는 실정이다. 줄기세포 연구의 발달로 인해 인류가 난치병 치료의 첫 장을 열었다고 하더라도 그 영향이 당장 보험사에 미친다고 할 수는 없다. 왜냐하면 앞으로 이러한 신기술이 실제 의료행위에 적용되기 위해서는 여러 단계의 안정화 작업과 임상시험이 필요한데 이러한 작업이 기술적으로 어렵고 그 시간도 만만치 않게 걸리기 때문이다. 또한, 보험사의 보장은 크게 사망/수술/입원/암/기타보장으로 구별할 수 있는데, 줄기세포 연구의 발달과 관련이 있는 보장이 제한되어 있어 보험사에 미치는 영향이 당장 우려할 만한 수준이라 할 수 없다. 하지만 만약 치료용 줄기세포 배양으로 인한 장기 기관의 이식이나 손상세포의 대체 등과 같은 의학신기술의 예상 외로 급격하게 발전한다면 보험사의 Risk 관리에 상당한 저해요인으로 작용할 것으로 판단된다. 특히 진단 입원 수술로 대표되는 생존보장에 대한 사차 Risk 및 사차손의 급증이나 역선택 증가는 보험사의 경영수지 악화를 유발하여 보험산업 전반에 위험으로 작용할 수도 있다. 따라서, 장기적인 안목으로 업계 공동의 대응이 필요하고, 각 사에서도 상품개발, 언더라이팅, 지급심사 간의 긴밀한 협조가 요구된다. 생명보험산업의 Risk 관리는 기존의 시장환경에 영향을 받는 비차, 이차중심에서 보험회사가 어느 정도 관리를 통해 적정규모를 유지할 수 있는 사차로 그 중심축이 이동하고 있다. 보험산업이 계속 활력을 갖고 성장하기 위해서는 체계적인 Risk나 관리가 핵심일 것이며, 보험사의 사차 Risk의 중요성이 더욱 커져 가고 있는 현실에서 거시적으로 의학신기술 발달 등 위험요인에 대해 미리 분석하고 이에 대한 대비책 마련이 필요할 것으로 판단된다.

• Development and Reproduction
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• v.12 no.1
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• pp.1-8
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• 2008

Specific protocols to increase the differentiation of neuronal cells from embryonic stem (ES) cells have been well established, such as retinoic acid induction and lineage selection of neuronal cells. For the neuropathological studies, ES-derived neurons (ES neurons) must show normal physiological characteristics related to cell death and survival and should be maintained in vitro for a sufficient time to show insults-specific cell death without spontaneous death. When mouse ES cells were plated onto astrocytes monolayer after retinoic acid induction, most ES cells differentiated into neuronal cells, which were confirmed by the presence of specific neuronal markers, and the cultures were viable for at least four weeks. When these cultures were examined for vulnerability to glutamate excitotoxicity, ES neurons were vulnerable to excitotoxic insults mediated by agonist-specific receptors. The vulnerability to excitotoxic death increased with developmental age of ES neurons in vitro. Specific receptors for Neurotrophin and GDNF family ligands were present in ES neurons. GDNF and NT-3 could modulate the survival and excitotoxic vulnerability of ES neurons. The vulnerability and resistance to toxic insults, which are essential requirements of model culture systems for neuropathological studies, make ES neurons to a useful model culture system. Especially ES cell are highly amenable to genetic modification unlikely to primary neuronal cells, which will give us a chance to answer more complicated neurophysiological questions. Recently there was an outstanding attempt to explore the cellular toxicity using human ES cells (Schrattenholz & Klemm, 2007) and it suggested that ES cells could be a new model system for neurophysiological studies soon and go further a large-scale screening system for pharmacological compounds in the future.