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dc.contributor.advisor이승진-
dc.contributor.authorPramila Katila-
dc.creatorPramila Katila-
dc.date.accessioned2016-08-26T04:08:44Z-
dc.date.available2016-08-26T04:08:44Z-
dc.date.issued2015-
dc.identifier.otherOAK-000000111510-
dc.identifier.urihttps://dspace.ewha.ac.kr/handle/2015.oak/212248-
dc.identifier.urihttp://dcollection.ewha.ac.kr/jsp/common/DcLoOrgPer.jsp?sItemId=000000111510-
dc.description.abstractThe utility of hepatic stem cells (HSCs) could provide a novel solution to the severe shortage of human donor livers, for treatment of liver related disease, due to their ability to proliferate and differentiate into functional hepatocytes. However, morphology, phenotype, successful isolation and culture of HSC require much investigation. Several studies have suggested the presence of stem cells in the adult normal human liver; however, a population with stem cell properties has not been isolated. Here, we describe a fibrin matrix-supported three-dimensional (3-D) organ culture system for liver tissue. This culture system included the use of the fibrin hydrogel coupled with dynamic culture conditions. This culture system allowed HSCs outgrowth into the hydrogel to be robustly supported, while maintaining the structural integrity of liver tissue during in vitro culture. The outgrown cells (OCs) were successfully recovered from fibrin by urokinase treatment and propagated. These OCs showed the characteristics of mesenchymal stem cells (MSCs), adherence to plastic, multipotency and cell surface molecule expressions. HSCs expressed the MSCs markers such as CD29, CD73, CD44, CD90, and CD105. HSCs were also able to undergo osteogenic, adipogenic and hepatogenic differentiation when cultured in the appropriate differentiation media. Our results suggest that the endogenous HSCs present in the rat liver have MSCs like properties. In addition, HSCs when cultured with perfluorodecalin (PFD) showed greater cell viability and proliferation. Our results indicate that this organ culture of liver tissue in 3-D supported fibrin matrix can be a unique strategy for isolating endogenous hepatic stem cell having MSCs like property;증식 및 분화능을 지닌 간 줄기세포를 이용은 심각한 간 기증자의 부족에 대한 새로운 대책이 될 수 있다. 그러나 간 줄기세포의 형태, 표현형 및 분리 및 배양법에 관하여 더욱 연구되어야 한다. 여러 연구에서, 성인의 정상 간에 줄기세포가 존재함이 시사되었으나, 줄기세포의 특성을 갖고 있는 세포군이 분리된 적은 없었다. 본 연구에서는, 간 조직 피브린 기반 삼차원 조직 배양 시스템에 관하여 보고자 한다. 이 배양 시스템은 다이나믹 배양 조건을 수반한 피브린 하이드로젤이 사용되었다. 이 in vitro 시스템 상에서, 간 조직은 형태를 그대로 유지하면서 조직으로부터 하이드로젤 내로 간 줄기세포가 자라 나왔다(outgrown). 조직으로부터 자라나온 세포를 유로키나아제를 처리하여 피브린으로부터 성공적으로 분리하여 배양했다. 또한, 이 세포는 중간엽 줄기세포의 특성을 지니고, 세포 배양디쉬에 붙어자라면서 다분화 능 및 세포 표면 마커들을 발현하였다. 간 줄기세포는 중간엽 줄기세포의 마커인 CD29, CD73, CD44, CD90 및 CD105를 발현하였으며, 각각의 분화배지를 이용하여 분화시켰을 때, 골, 지방 및 간 세포로의 분화능을 보였다. 이 결과는 mouse의 간로부터 분리한 간 줄기세포가 중간엽 줄기세포의 특성을 지녔다는 것을 알 수 있었다. 또한, 이 세포를 perfluorodecalin (PFD) 조건에서 배양하였을 때, 더 높은 세포 viability와 증식능을 보였다. 이 결과는, 삼차원 피브린 매트릭스가 중간엽 줄기세포의 특성을 지니면서 내재 간 줄기세포 분리의 새로운 전략이 됨을 보여준다.-
dc.description.tableofcontents1. Introduction 1 2. Materials and Methods 7 2.1. Materials 7 2.2. 3-D organ culture of liver tissue and isolation of outgrown cells (OCs) in fibrin matrix 7 2.3. Culture Media 10 2.4. Flow Cytometry Analysis of Subcultured OCs 10 2.5. In vitro differentiation into adipogenic, osteogenic, and hepatogenic lineages 11 2.6. Preparation of Perfluorodecalin (PFD) 12 2.7. Preparation of PFD Emulsion 12 2.8. Proliferation assay of HSCs 13 3. Results 14 3.1. Establishment of hydrogel-supported 3-D organ culture of liver tissue 14 3.2. In vitro expansion of HSCs under monolayer culture condition 16 3.3. Immunophenotypic characteristics of in vitro expanded Hepatic Stem Cells 17 3.4. In vitro expanded HSCs show the developmental capacity of MSCs 18 3.5. In vitro differentiation Potential into hepatocyte of in vitro expanded HSC 19 3.6. Preparation of PFD emulsion 20 3.7. Effects of perfluorodecalin (PFD) on Hepatic Stem Cells proliferation 20 4. Discussion 22 5. Conclusion 26 References 27 Abstract in Korean 33-
dc.formatapplication/pdf-
dc.format.extent1224639 bytes-
dc.languageeng-
dc.publisher이화여자대학교 대학원-
dc.subject.ddc600-
dc.title3-D fibrin supported organ culture and isolation of endogenous hepatic stem cells having MSCs-like property-
dc.typeMaster's Thesis-
dc.format.pagevi, 34 p.-
dc.contributor.examiner양영일-
dc.contributor.examiner이혁진-
dc.contributor.examiner이승진-
dc.identifier.thesisdegreeMaster-
dc.identifier.major대학원 약학과-
dc.date.awarded2015. 2-
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일반대학원 > 약학과 > Theses_Master
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