초록 ▼

기술개발내용: COL1A2 프로모터-루시퍼라아제를 발현하는 인간 fibroblast 세포주의 개발
1. stable cell line 제작에 필요한 stable cell용 리포더 벡터 완성(COL1A2 프로모터+루시퍼라아제 유전자+ selection marker)
2. 완성된 벡터를 인간 fibroblast 세포에 형질전환용 vehicle를 이용해 삽입함
3. G418 항생제를 이용, COL1A2 프로모터와 루시퍼라아제 유전자를 보유하는 세포를 선별함.
4. 선별된 세포를 콜라겐 합성에 관여하는 물질을 이용해

기술개발내용: COL1A2 프로모터-루시퍼라아제를 발현하는 인간 fibroblast 세포주의 개발
1. stable cell line 제작에 필요한 stable cell용 리포더 벡터 완성(COL1A2 프로모터+루시퍼라아제 유전자+ selection marker)
2. 완성된 벡터를 인간 fibroblast 세포에 형질전환용 vehicle를 이용해 삽입함
3. G418 항생제를 이용, COL1A2 프로모터와 루시퍼라아제 유전자를 보유하는 세포를 선별함.
4. 선별된 세포를 콜라겐 합성에 관여하는 물질을 이용해 기능 테스트를 수행, 최적의 세포주를 최종 선발함.
5. 개발된 세포주를 이용, 최소 1종이상의 주름개선 물질을 선발함:
- 2종선발 (동백오일, 인삼추출물)
- 논문투고(인삼추출물관련, Journal of Ethnopharmacology, 2006년)
- 논문 준비중(스크리닝시스템구축, Analytical Biochemistry)
- 특허 출원완료 (제목: 인삼류의 생리활성물질 증진방법, 출원번호: 10-2006-0025865, 출원일 2006년 3월 21일)

Abstract ▼

The collagens are the major structural component of the skin and represent a large family of extracellular proteins that impart specific physical properties to tissues in addition to Playing important roles during morphogenesis and growth and in tissue homeostasis and repair. With increasing age, co

The collagens are the major structural component of the skin and represent a large family of extracellular proteins that impart specific physical properties to tissues in addition to Playing important roles during morphogenesis and growth and in tissue homeostasis and repair. With increasing age, collagen levels are reduced.
Alterations incollagen have been considered to be s cause of skin aging, and are observed in naturally aged and photoaged skin.
Type I collagen consists of two alphalchains and one alpha2 chain that are produced by two fairly large genes that reside on different chromosomes in both the human and the mouse genome. And it is the most abundantly expressed member of the collagen gens, there by playing a significantly role in maintaining homeostasis.
Transforming growth factor-$\beta$ (TGF-$\beta$) is well known to be the principal factor that induces type I collagen gene expression and leads to tissue fibrosis. The Smads have been identified ad intracellular mediators of the signal transduction pathways of TGF-$\beta$ superfamily member to function down stream from the serine/threonine kinase receptors of the TGF-13 family to transducersignal to the nucleus. Smad3 promotes collagen gene (COL1A2) activation in human glomerular mesangial cells, potentially contributing to glomerularosis and Sp1 binding is necessary for TGF-$\beta$1-induced type I collagen mRAN expression. Sp1 and Smad proteins form complexes and their synergy plays an important role in mediating TGF-$\beta$1-induced type I collagen expression in human mesangial cells. Sp1 binding is required for Smad3 stimulation of COL1A2 promoter activity and is essential for the human glomerular mesangial cell responsiveness to TGF-$\beta$ 1. Sp 1 and Smad proteins form complexes, and their synergistic cooperation plays an important role in mediating TGF-$\beta$1-induced collagen expression. Therefore, monitoring COL1A2 promoter activity in human skin cells could potentially provide an appropriate means for screening of anti-wrinkle agents.
We established stable human normal fibroblast (HNF) cells transfected pLuc-378COL1A2 plasmid an pCl-neo plasmid for quantitative measurement of COL1A2 promoter activity by various synthetic chemicals or natural products. The pLuc-378COL1A2 plasmid expresses the luciferase gene in response to the level of COL1A2 promoter activity. And the pCI-neo plasmid is the dominant selective marker of neomycin resistance. Human normal fibroblast cell transfectants released the luciferase enzyme into the culture medium. We have further tested whether this assay system could be used for the quantitative measurement of the level of COL1A2 promoter activity induced by TGF-$\beta$1.
We report that this HNF cell-based reporter system in response to COL1A2 promoter activity could be used for the screening of anti-wrinkle agents.

목차 Contents

• 표지...1
• 제출문...2
• 보고서 요약서...3
• 요약문...4
• SUMMARY...6
• CONTENTS...17
• 목차...18
• 제 1 장 연구개발과제의 개요...19
• 제 2 장 국내외 기술개발 현황...21
• 제 3 장 연구개발수행 내용 및 결과...21
• 제 4 장 목표달성도 및 관련분야에의 기여도...30
• 제 5 장 연구개발결과의 활용계획...31
• 제 6 장 연구개발과정에서 수집한 해외과학기술정보...33
• 제 7 장 참고문헌...33
• 제 8 장 첨부서류...35