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케라틴을 함유한 PLGA지지체에서 OECs의 증식 및 표현형 분석
전나리 ( Na Ri Jeon ),조은혜 ( Eun Hye Jo ),박현진 ( Hyun Jin Park ),( Jin San Choi ),( Sang Jin Lee ),( Mark Van Dyke ),( James J Yoo ),( James J Lee ),이동원 ( Dong Won Lee ),이종문 ( John M Rhee ),강길선 ( Gil Son K 한국조직공학·재생의학회 2009 조직공학과 재생의학 Vol.6 No.4
Keratin that affects to cell proliferation and adheison has been used to clinical trial as natural material. In this study, we evaluated keratin/PLGA scaffolds on adhesion, proliferation and phenotype of olfactory ensheathing cells(OECs). OECs were isolated from olfactory bulb of Fischer rat. We fabricated keratin/PLGA scaffold using 0, 10, 20, and 50 wt% of keratin by the solvent casting/salt leaching method. We confirmed morphology of cellular adhesion by scanning electron microscope(SEM). Cell proliferation activity was measured by MTT assay. RT-PCR was conducted to confirm mRNA expression of NF, S100 and P75 for OECs marker. These results suggest that keratin provides suitable surface to OECs and keratin of suitable content affects to improve cell adhesion and phenotype maintenance.
케라틴/PLGA 지지체가 골수간엽줄기세포의 신경분화에 미치는 영향
오아영 ( A Young Oh ),김순희 ( Soon Hee Kim ),김초민 ( Cho Min Kim ),정수현 ( Su Hyun Jung ),전나리 ( Na Ri Jeon ),홍현혜 ( Hyun Hye Hong ),( Sang Jin Lee ),( Mark Van Dyke ),( James J Yoo ),이동원 ( Dong Won Lee ),이종문 ( 한국조직공학·재생의학회 2009 조직공학과 재생의학 Vol.6 No.1
Bone marrow stromal cells(BMSCs) exhibit multiple traits of a stem cell population, and they can expand many times in vitro and be induced to differentiate into multiple cell types. Keratin is the major structural fibrous protein providing outer covering such as wool, hair, and nail. We think keratins are useful as natural biomaterial. We examined the effect of Keratin/PLGA scaffold on the neural differentiation of rat BMSCs. We developed the keratin loaded poly(L-lactide-co-glycolide)(PLGA) scaffolds. Keratin/PLGA 0, 20 and 50 wt% scaffolds were prepared by solvent casting/salt leaching method. BMSCs were harvested from the femurs of adult female Fischer rat. These cells were seeded in prepared Keratin/PLGA scaffold and cultured in Medium 50 ml DMEM, 2 %DMSO, 200 ?M BHA, 25 ?M valproic acid, 10 ?M forskolin, 1 ?M hydro-cotisone, 5?g/ml insulin for 5 day. The effect of Keratin/PLGA scaffold on the neural differentiation of rat BMSCs were assessed in culture using the MTT assay and RT-PCR was conducted to confirm mRNA expression of NSE and Nf for neural marker. We confirmed that effect of Keratin/PLGA scaffold on the neural differentiation of rat BMSCs. According to our results, 20 wt% keratin/ PLGA scaffold have good cell compatibility and the expression of neural markers was higher. In conclusion, Keratin/ PLGA scaffold, on which neural differentiation of rat BMSCs was possible.
케라틴을 함유한 PLGA 지지체가 슈반세포의 증식 및 특성 유지에 미치는 영향
오아영 ( A Young Oh ),김순희 ( Soon Hee Kim ),정수현 ( Su Hyun Jung ),홍현혜 ( Hyun Hye Hong ),전나리 ( Na Ri Jeon ),( Sang Jin Lee ),( Mark Van Dyke ),( James J. Yoo ),이종문 ( John M. Rhee ),강길선 ( Gilson Khang ) 한국조직공학·재생의학회 2008 조직공학과 재생의학 Vol.5 No.4
Keratin is the major structural fibrous protein such as wool, hair, and nail and is useful as natural protein. Schwann cells(SCs) can stimulate tissue sparing and enhance outgrowth of both intact and lesion axons. We were prepared keratin/PLGA(0, 20 and 50 wt%) scaffolds by solvent casting/salt leaching method. Cellular viability and proliferation were assayed by MTT test. Morphology of cellular adhesion were confirmed by scanning electron microscope(SEM). SCs specific protein was assessed staining by s-100 for SCs marker and RT-PCR was conducted to confirm mRNA expression of NF, P-75 and s-100 for SCs marker. In MTT assay results, cell viability in scaffolds impregnated 20 wt% of keratin were higher than other scaffolds. SEM exhibited normal spindle-shaped morphology on 20 wt% of keratin. SC specific mRNA expression and protein could not be observed in scaffold containing 50 wt% of keratin. These results suggest that keratin provide suitable surface to neural cells and proper content affect on culture condition to improve cell adhesion and proliferation.
조직공학적 연골재생을 위한 In Vivo 환경에서의 케라틴/PLGA 지지체의 효과
정수현 ( Su Hyun Jung ),김순희 ( Soon Hee Kim ),양재찬 ( Jae Chan Yang ),홍현혜 ( Hyun Hye Hong ),김혜린 ( Hye Lin Kim ),김원 ( Won Kim ),손영숙 ( Young Sook Son ),( Sang Jin Lee ),( Mark Van Dyke ),( James J Yoo ),이종문 ( 한국조직공학·재생의학회 2009 조직공학과 재생의학 Vol.6 No.1
Articular cartilage that is difficult to recovery when damaged needs to tissue engineering. keratin are the intermediate filament proteins that form a dense meshwork of filaments throughout the of cells and generally expressed in particular pairs of type I and type II keratin proteins in a-specific and cellular differentiation-specific manner. In this study, we are developing an alternative approach that consists of generating chondrocytes anchored to poly(L-lactide-co-glycolide)(PLGA) scaffolds impregnated keratin(keratin/PLGA) using tissue-engineering principles. We prepared PLGA and keratin/PLGA scaffolds using solvent casting/salt leaching method. Chondrocytes were isolated from the articular cartilage of New Zealand white rabbit and cultured With DMEM/Ham`s F-12 supplemented with 10 % FBS, 1 % penicillin streptomycin, 200 mM L-glutamin, 50 ?g/ml of ascorbic acid and 15 mM HEPES buffer 1M. After 2weeks of cell seeding, we implanted keratin /PLGA scaffolds on the back of nude mice. Morphological observation, histology, biological assay for collagen and sGAG, and PCR were performed at each time point 1, 2, 3 and 6 weeks. The cell viability and the quantity of collagen and sGAG were better keratin/PLGA scaffolds than PLGA scaffolds. Specific mRNA, type II and type I collagen, for chondrocyte expressed significantly highly in keratin/PLGA scaffold. keratin/PLGA scaffold promotes in vivo chondrocyte of rabbit articular chondrocytes. This study suggests that keratin/PLGA scaffold may serve as a potential cell delivery vehicle and a structural basis for in vivo tissue engineered articular cartilage.
In vitro상에서 연골재생을 위한 케라틴/PLGA 지지체의 효과
홍현혜 ( Hyun Hye Hong ),김순희 ( Soon Hee Kim ),오아영 ( A Young Oh ),전나리 ( Na Ri Jeon ),정수현 ( Su Hyun Jung ),( Sang Jin Lee ),( Mark Van Dyke ),( James J. Yoo ),손영숙 ( Youngsook Son ),이종문 ( John M. Rhee ),강길선 한국조직공학·재생의학회 2008 조직공학과 재생의학 Vol.5 No.4
Synthetic and naturally derived biodegradable polymers have been widely used to construct scaffolds for cartilage tissue engineering. We developed the keratin loaded poly(L-lactide-co-glycolide)(PLGA) scaffolds(keratin/ PLGA) to utilize as highly functional scaffolds for tissue engineering by improving hydrophobicity and cell compatibility of the polymer scaffolds. Keratin as natural protein is the major structural fibrous protein providing outer covering such as wool, hair, and nail. Keratin/PLGA scaffolds were prepared by casting/salt leaching method. Cell proliferation activity was measured via MTT assay. Scaffold mechanical strength, histology, gene expression, sulphated- glycosaminoglycan(sGAG) and collagen contents analyses were performed to elucidate in vitro cartilage development and the deposition of cartilage-specific extracellular matrices. We concluded in vitro chondrogenesis of articular chondrocytes was improved in PLGA scaffolds containing keratin.