백렴 에틸 아세테이트 층의 항산화 활성과 Matrix Metalloproteinase-1 발현 저해효과

Effects of Ethyl Acetate Fraction from Melothria Heterophylla on Antioxidant Activity and Matrix Metalloproteinase-1 Expression in Ultraviolet A-irradiated Human Dermal Fibroblasts

  • 조영호 (한불화장품(주) 기술연구소) ;
  • 김진희 (한불화장품(주) 기술연구소) ;
  • 심관섭 (한불화장품(주) 기술연구소) ;
  • 이동환 (한불화장품(주) 기술연구소) ;
  • 이범천 (한불화장품(주) 기술연구소) ;
  • 표형배 (한불화장품(주) 기술연구소)
  • 발행 : 2005.03.01

초록

자외선은 콜라겐 분해와 같은 피부 결합조직에서 특이적인 변화를 유발한다. 세포외 기질(extracelluar matrix)내에서의 많은 변형들은 기질 금속 단백질 분해효소(matrix metalloproteinases)에 의해 매개된다. 본 연구에서는 천연물 유래의 새로운 노화방지소재를 개발하기 위해 백렴 추출물의 용매별 분획들의 항산화 활성을 검색하고, 그 중에서 활성이 가장 높게 나타난 에틸아세테이트 층에 대해 MMP-1 활성 및 human dermal fibroblasts에서 자외선에 의한 MMP-1 발현에 미치는 영향을 측정하였다. 백렴 추출물의 에틸 아세테이트 층은 MMP-1의 활성을 농도 의존적으로 저해하였다($IC_{50}=9{\mu}g/mL$). 또한, 자외선에 의해 증가되는 MMP-1의 발현이 에틸 아세테이트 층을 $100{\mu}g/mL$ 처리한 경우 약 $90\%$ 정도 저해되었다. 반면에 MMP-1 mRNA의 발현에는 별다른 영향을 미치지 않는 것으로 나타났다. 따라서 백렴 에틸 아세테이트 층은 MMP-1의 발현을 단백질 수준에서만 저해함을 알 수 있다. 결론적으로 백렴 에틸 아세테이트 층은 자외선에 의해 손상된 피부를 보호할 수 있는 새로운 노화방지 소재로 이용될 수 있다.

UV irradiation leads to distinct changes in skin connective tissue, which is degradation of collagen. Many of these alterations in the extracellular matrix are mediated by matrix metalloproteinases. In this study, to develop a new anti-aging agent, we screened the antioxidant activity of solvent fractions from ethanolic extract of Melothria Heterophylla. Among the four solvent fractions tested, the EtOAc fraction exhibited the highest antioxidant activity. It was investigated the inhibitory effect of the EtOAc fraction on the expression and activity of MMP-1 in UVA-irradiated human dermal fibroblasts. The EtOAc fraction inhibited the activity of MMP-1 in a dose dependent manner with the $IC_{50}$ values of $9{\mu}g/mL$. Also, UVA-induced MMP-1 expression was reduced about $90\%$ by $100{\mu}g/mL$ of the EtOAc fraction but MMP-1 mRNA expression was not inhibited. Therefore, we conclude that the EtOAc fraction significantly inhibits MMP-1 expression at the protein level. From these results, we suggest that the EtOAc fraction from M. heterophylla could be used as a new anti-aging agent for the photo-damaged skin.

키워드

참고문헌

  1. B. A. Gilchrest, Skin aging and photoaging: an overview, J. Am Amd. Dermaiol., 21, 610 (1989) https://doi.org/10.1016/S0190-9622(89)70227-9
  2. E. Cadenas, Biochemistry of oxygen toxicity, Annu. Rev. Biochem., 58, 79 (1989) https://doi.org/10.1146/annurev.bi.58.070189.000455
  3. K. J. Davies, Protein damage and degradation by oxygen radicals, J. Biol. Chem., 262, 9895 (1987)
  4. C. Huang, W. Y. Ma, M. I. Dawson, M. Rincon, R. A. Flavell, and Z. Dong, Blocking activator protein1 activity, but not activation retinoic acid response element, is required for the antitumor promotion effect of retinoic acid, Proc. Natl. Acad. Sci. USA, 94, 5826 (1997)
  5. S. Kondo, The roles of cytokines in photoaging, J. Dermatol. Sci., 23, S30 (2000) https://doi.org/10.1016/S0923-1811(99)00076-6
  6. Y. Wang, A. R. Johnson, Q. Z. Ye, and R. D. Dyer, Catalytic activities and substrate specificity of the human membrane type 4 matrix metalloproteinase catalytic domain, J. Biol. Chem., 274, 33043 (1999) https://doi.org/10.1074/jbc.274.46.33043
  7. P. Brenneisen, H. Sies, and K Scharffetter-Kochanek, Ultraviolet-B irradiation and matrix rnetalloproteinases: from induction via signaling to initial events, Ann. N.Y. Acad. Sci., 973, 31 (2002) https://doi.org/10.1111/j.1749-6632.2002.tb04602.x
  8. J. H. Park, The encyclopedia of Chinese crude drugs, 310, Shinilbooks Publications, Seoul (2002)
  9. Y. H. Cho, G. S. Sim, J. H. Kim, S. M. Park, B. C. Lee, H. B. Pyo, Y. P. Yun, and H. D. Park, Effects of Melothria heterophylla Extract on Expression of Matrix Metalloproteinase-1 in Human Skin Fibroblasts, Yakhak Hoeji, 48, 358 (2004)
  10. M. S. Blois, Antioxidant determinations by the use of a stable free radical, Nature, 181, 1199 (1958) https://doi.org/10.1038/181958a0
  11. K. Furuno, T. Akasako, and N. Sugihara, The contribution of the pyrogallol moiety to the superoxide radical scavenging activity of flavonoids, Biol. Pharm. Bull., 25, 19 (2002) https://doi.org/10.1248/bpb.25.19
  12. T. Mosmann, Rapid colorimetric assay for the cellular growth and survival: application to proliferation and cytotoxic assay, J. Immunol. Methods., 65, 55 (1983) https://doi.org/10.1016/0022-1759(83)90303-4
  13. S. E. Dunsmore, J. S. Rubin, S. O. Kovacs, M. Chedid, W. C. Parks, and H. G. Welgus, Mechanisms of hepatocyte growth factor stimulation of keratinocyte metalloproteinase production, J. Biol. Chem., 271, 24576 (1996) https://doi.org/10.1074/jbc.271.40.24576
  14. P. Kuppusamy and J. L. Zweier, Characterization of free radical generation by xanthine oxidase, J. Biol. Chem., 264, 9880 (1989)
  15. J. H. Chun, S. W. Kang, J. Varani, J. Lin, G. J. Fisher, and J. J. Voorhees, Decreased extracellularsignal regulated kinase and increased stress-activated MAP kinase activities in aged human skin in vivo, J. Invest. Dermatol., 115, 177 (2000) https://doi.org/10.1046/j.1523-1747.2000.00009.x
  16. G. J. Fisher, H. S. Talwar, J. Lin, P. Lin, P. McPhillips, Z. Q. Wang, X. Li, Y. Wan, S. W. Kang, and J. J. Voorhees, Retinoic acid inhibits induction of c- jun protein by ultraviolet radiation that occurs subsequent to activation of mitogenactivated protein kinase pathways in human skin in vivo, J. Clin. Invest., 101, 1432 (1998) https://doi.org/10.1172/JCI2153