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천연재료가 함유된 조직공학용 지지체에 대한 세포독성 분석법의 비교
백미옥 ( Mi Ock Baek ),김순희 ( Soon Hee Kim ),소정원 ( Jung Won So ),노혜원 ( Hye Won Roh ),이나리 ( Na Ri Lee ),김문석 ( Moon Suk Kim ),유규하 ( Gyu Ha Ryu ),조양하 ( Yang Ha Cho ),이승진 ( Seung Jin Lee ),강길선 ( Gilson Khang 한국조직공학·재생의학회 2007 조직공학과 재생의학 Vol.4 No.4
Cytotoxicity measurements on materials are extremely important in tissue engineering field. The objective of this study is to measure and compare the cytotoxicity of scaffold for tissue engineered using 3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyl-tetrazolium bromide(MTT), 2-(methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2-4- disulfophenyl)-H-tetrazolium(WST-8) and Alamar Blue cytotoxicity assay on cytotoxixity of scaffold for tissue engineering. In order to evaluate the sensitivity of rat bone marrow stromal cell(rBMSC) for cytotoxicity assays, a series of two-fold dilution of rBMSC were plated in 96-wells. rBMSC suspension was seeded on poly(lactide-coglycolide) (PLGA) scaffold and then cytotoxicity was measured in vitro by MTT, WST-8 and alamar blue assay. The MTT assay was more sensitive than WST-8, alamarblue assay for rBMSC. rBMSC suspension seeded on various content of demineralized bon particle(DBP) impregnated PLGA scaffold was measured to confirm on affect to natural material by MTT assay. In the MTT assay results, optical density was influenced by content of DBP. Finally, cytotoxicity of DBP/PLGA scaffold was assayed using modified method of ASTM F813-07. We confirmed that cell viability was significantly high in PLGA scaffold including DBP of 20 and 40% compared to other scaffolds. This report may be useful as guideline in tissue engineered scaffolds.
탈미네랄화된 골분용액을 함침시킨 PLGA 지지체의 제조 및 특성
백미옥 ( Mi Ock Baek ),김순희 ( Soon Hee Kim ),소정원 ( Jung Won So ),임지예 ( Ji Ye Lim ),최진희 ( Jin Hee Choi ),이종문 ( John M. Rhee ),이해방 ( Hai Bang Lee ),강길선 ( Gilson Khang ) 한국조직공학·재생의학회 2008 조직공학과 재생의학 Vol.5 No.2
We developed the synthetic/natural hybrid scaffolds with poly (lactide-co-glycolide)(PLGA) and solution using DBP(DB solution) for tissue engineering. Composite scaffolds of PLGA scaffold penetrated DB solution (PLGA-pen-DBP) were manufactured by simple method that PLGA scaffolds manufactured by solvent casting/salt leaching method soaked in DB solution. We evaluated pore structure, wettability, water uptake ability and cell viability of two types of scaffolds. Through SEM and FT-IR, it was confirmed that DB solution was put into the pores of PLGA scaffold. Also, the effects of the impregnation of DB solution on PLGA scaffolds were characterized through water uptake ability and cell viability. It seemed that DBP provided proper pore to PLGA scaffold and improved water absorption. A novel scaffold, which PLGA scaffold penetrated with DB solution, might be suitable to apply as tissue engineered scaffold.
ASTM F2027 조직공학적 의료용제품(TEMPs)을 위한 기질의 특성결정 및 테스트를 위한 표준 지침서
백미옥 ( Mi Ock Baek ),소정원 ( Jung Won So ),김순희 ( Soon Hee Kim ),노혜원 ( Hye Won Roh ),이나리 ( Na Ri Lee ),김문석 ( Moon Suk Kim ),유규하 ( Gyu Ha Ryu2 ),조양하 ( Yang Ha Cho ),이승진 ( Seung Jin Lee ),강길선 ( Gilson Khang 한국조직공학·재생의학회 2007 조직공학과 재생의학 Vol.4 No.4
Tissue engineering medical products are those protocols and products developed for use in the human body as biological substitutes to restore, maintain, or improve tissue function. The purpose of this standard is to locate relevant existing guideline and test methods and to provide guidance for interim use of materials for which a standard does not exist. The standard may be use as guideline in tissue engineered research.
PLGA 지지체에 함유된 DBP 함량이 줄기세포의 골분화에 끼치는 영향에 대한 in vitro 테스트
백미옥 ( Mi Ock Baek ),김순희 ( Soon Hee Kim ),소정원 ( Jung Won So ),임지예 ( Ji Ye Lim ),노혜원 ( Hye Won Roh ),이나리 ( Na Ri Lee ),유규하 ( Gyu Ha Ryu ),조양하 ( Yang Ha Cho ),이승진 ( Seung Jin Lee ),강길선 ( Gil Son Khang ) 한국조직공학과 재생의학회 2008 조직공학과 재생의학 Vol.5 No.1
Demineralized bone particles(DBP) exhibit osteoinductive properties and are very useful for bone formation and other orthopeadic applications. In this study, we investigated the osteogenic capabilities of rat bone marrow stromal cells(rBMSCs) in PLGA scaffold impregnated with various DBP contents. DBP-loaded PLGA scaffold (10, 20, 40, 80 % w/w) were prepared by solvent casting/salt leaching method. rBMSCs were seeded in PLGA and DBP/PLGA scaffolds, and then cellular viability and proliferation were assayed by water-soluble tetrazolium salt(WST-8) assay. Osteoblast differentiation of cells were analyzed by alkaline phosphatase(ALP) activity. Finally, reverse transcript polymerase chain reaction(RT-PCR) was assessed to measure mRNA expression of the genes (ALP, osteocalcine and type I collagen) of osteogenic marker in rBMSCs depending on various content of DBP. These results could demonstrated that presence of DBP in PLGA might play important roles in osteoblast differentiation of rBMSCs.
ASTM F813-07 의료용디바이스를 위한 생체재료의 직접촉 세포배양측정에 대한 표준지침서
김순희 ( Soon Hee Kim ),백미옥 ( Mi Ock Baek ),소정원 ( Jung Won So ),노혜원 ( Hye Won Roh ),이나리 ( Na Ri Lee ),김문석 ( Moon Suk Kim ),유규하 ( Gyu Ha Ryu2,),조양하 ( Yang Ha Cho ),이승진 ( Seung Jin Lee ),강길선 ( Gilson Khang 한국조직공학·재생의학회 2007 조직공학과 재생의학 Vol.4 No.4
We provided standard practice for direct contact cell culture evaluation of materials for medical devices in this report. This practice introduced a reference method of direct contact cell culture testing. This method of direct contact cell culture may be used in the construction of medical materials and devices for test the cytotoxic potential of materials. Researcher for medical devices could use this practice to compare other cytotoxicity tests or directly to evaluate materials. Cell or materials could be sometimes modified depending on use in offered extend of this practice.
PLGA 담체의 제조방법에 따른 골수간엽줄기세포의 골분화 유도
소정원 ( Jeong Won So ),김순희 ( Soon Hee Kim ),백미옥 ( Mi Ock Baek ),임지예 ( Ji Ye Lim ),노혜원 ( Hye Won Roh ),이나리 ( Na Ri Lee ),유규하 ( Gyu Ha Ryu ),조양하 ( Yang Ha Cho ),이승진 ( Seung Jin Lee ),강길선 ( Gilson Khang ) 한국조직공학·재생의학회 2008 조직공학과 재생의학 Vol.5 No.1
A key component in tissue engineering for bone regeneration is the scaffold that serves as a template for cell interaction and the formation of bone-extracellular matrix to provide structural support to the newly formed tissue. In this study, poly(L-Lactide-co-glicolide)(PLGA) scaffolds were prepared by solvent casting/salt-leaching, ice particle-leaching and gas-forming/salt-leaching. Cellular viability and proliferation were assayed by 3-(4,5-dimethylthiazole- 2-yl)-2,5-diphenyltetrazolium-bromide(MTT) test. Osteogenic differential cells were analyzed by ALP activity. Through RT-PCR analysis, we observed ALP and osteocalcin m RNA expression. Scaffold with BMSCs were implanted into the back of athymic nude mouse to observe in vivo behavior. Thin section were cut from paraffin embedded tissues and histological sections were stained by trichrome, immunohistochemical, von kossa staining. The cell proliferation and ALP activity of BMSCs in the scafoolds was the highest in the scaffolds using solvent casting/salt-leaching method. In RT-PCR results, in the all type of scaffolds observed specific gene expression of ALP, osteocalcin. By histological staining, we could observed collagen and calcification region in scaffolds by solvent casting/salt leaching method. Thus Solvent casting/salt-leaching method provided highly porous structure with good interconnections between each pores, which can support the surface of cell proliferation, differentiation. This result indicates that solvent casting/salt-leaching is useful for osteogenesis of BMSCs.
F2211-04 생체조직공학적 의료제품(TEMPs)에 대한 표준 분류
소정원 ( Jung Won So ),김순희 ( Soon Hee Kim ),백미옥 ( Mi Ock Baek ),노혜원 ( Hye Won Roh ),이나리 ( Na Ri Lee ),유규하 ( Gyu Ha Ryu ),조양하 ( Yang Ha Cho ),이승진 ( Seung Jin Lee ),강길선 ( Gil Son Khang ) 한국조직공학과 재생의학회 2008 조직공학과 재생의학 Vol.5 No.1
Tissue engineered medical products (TEMPs) are those protocols and products developed for use in the human body as biological substitutes to restore, maintain, or improve tissue function. So TEMPs may achieve a therapeutic potential from cells, biomolecules, scaffolds, and other materials, and processed tissue and derivatives used in various combinations or alone. Since interactions may occur among the components used in TEMPs, new standard descriptions, test methods, and practices are needed to aid the evaluation of these interactions. Therefore this technical report the aspects of tissue engineered medical that will be developed as standards.
PLGA미립구의 크기에 따른 인간디스크세포의 증식 및 표현형 분석
소정원 ( Jeong Won So ),김순희 ( Soon Hee Kim ),백미옥 ( Mi Ock Baek ),임지예 ( Ji Ye Lim ),노혜원 ( Hye Won Roh ),이나리 ( Na Ri Lee ),김문석 ( Moon Suk Kim ),유규하 ( Gyu Ha Ryu ),조양하 ( Yang Ha Cho ),이승진 ( Seung Jin Lee ) 한국조직공학·재생의학회 2007 조직공학과 재생의학 Vol.4 No.4
The PLGA microspheres can create an environment conductive to promising candidate as an injectable system for tissue engineered intervertebral disc(IVD). The final goal of this study was to investigate the condition of PLGA microspheres as microcarriers for IVD cells. We prepared PLGA microspheres using oil-in-water(O/W) solvent evaporation method and investigated the size variation of the microspheres(180~425 μm). Human disc cells were used in a prospective study of attachment, proliferation, and gene expression within various size of PLGA microspheres. Mophology of cell attached on the microspheres was studied using scanning electron microscopy (SEM). Cellular behavior in response to various size of PLGA microspheres was analyzed using MTT assay and reverse transcriptase - polymerase chain reaction(RT-PCR). SEM showed that cells could be bridge between microspheres with elongated shapes. MTT assay revealed that the cells could be attach, proliferate and spread on PLGA microspheres, in particular on the ones having size below 250 μm. In RT-PCR results, we observed strongly specific gene expression of type collagen at more 250 μm microsphere. In conclusion, this result will be used to injectable system for disc regeneration.