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Tyrosine Hydroxylase 유전자가 주입된 인간 배아줄기세포의 체외 신경세포 분화
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  • Tyrosine Hydroxylase 유전자가 주입된 인간 배아줄기세포의 체외 신경세포 분화
  • In vitro Neural Cell Differentiation of Genetically Modified Human Embryonic Stem Cells Expressing Tyrosine Hydroxylase
저자명
신현아,김은영,이금실,조황윤,김용식,이원돈,박세필,임진호,Shin. Hyun-Ah,Kim. Eun-Young,Lee. Keum-Sil,Cho. Hwang-Yoon,Kim. Yong-Sik,Lee. Won-Don,Park. Se-P
간행물명
대한불임학회지
권/호정보
2004년|31권 1호|pp.67-74 (8 pages)
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이 논문은 한국과학기술정보연구원과 논문 연계를 통해 무료로 제공되는 원문입니다.
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기타언어초록

Objective: This study was to examine in vitro neural cell differentiation pattern of the genetically modified human embryonic stem cells expressing tyrosine hydroxylase (TH). Materials and Methods: Human embryonic stem (hES, MB03) cell was transfected with cDNAs cording for TH. Successful transfection was confirmed by western immunoblotting. Newly transfected cell line (TH#2/MB03) was induced to differentiate by two neurogenic factors retinoic acid (RA) and b-FGF. Exp. I) Upon differentiation using RA, embryoid bodies (EB, for 4 days) derived from TH#2/MB03 cells were exposed to RA ($10^{-6}M$)/AA ($5{ imes}10^{-2}mM$) for 4 days, and were allowed to differentiate in N2 medium for 7, 14 or 21 days. Exp. II) When b-FGF was used, neuronal precursor cells were expanded at the presence of b-FGF (10 ng/ml) for 6 days followed by a final differentiation in N2 medium for 7, 14 or 21 days. Neuron differentiation was examined by indirect immunocytochemistry using neuron markers (NF160 & NF200). Results: After 7 days in N2 medium, approximately 80% and 20% of the RA or b-FGF induced Th#2/MB03 cells were immunoreactive to anti-NF160 and anti-NF200 antibodies, respectively. As differentiation continued, NF200 in RA treated cells significantly increased to 73.0% on 14 days compared to that in b-FGF treated cells (53.0%, p<0.05), while the proportion of cells expressing NF160 was similarly decreased between two groups. However, throughout the differentiation, expression of TH was maintained ($sim$90%). HPLC analyses indicated the increased levels of L-DOPA in RA treated genetically modified hES cells with longer differentiation time. Conclusion: These results suggested that a genetically modified hES cells (TH#2/MB03) could be efficiently differentiated in vitro into mature neurons by RA induction method.