Identification of HSP70-inducing activity in Arnica montana extract and purification and characterization of HSP70-inducers.
BACKGROUND
The expression of heat shock proteins (HSPs), particularly HSP70, is receiving considerable attention in the field of cosmetics, particularly given our recent report that ultraviolet-induced melanin production, skin damage and wrinkle formation were all suppressed in transgenic mice expressing HSP70.


OBJECTIVE
In the present study, we searched for HSP70-inducers from a library of herbal extracts that have already been approved as quasi-pharmaceutical products in Japan. We selected an ethanol extract of Arnica montana (A. montana), based on its high HSP70-inducing activity and low cytotoxicity.


METHODS
Cell viability was determined by MTT method and expression of HPS70 was monitored by immunoblotting analysis.


RESULTS
From the extract, we purified and identified eight sesquiterpene lactones (AM1-8) as HSP70-inducers, among which AM-2 (helenalin 2-methylbutyrate) was selected due to its good HSP70-inducing properties and low cytotoxicity. Treatment of cultured mouse melanoma cells with AM-2 or A. montana extract up-regulated the expression of HSP70 in a dose-dependent manner. This treatment also activated heat shock factor-1, a transcription factor for hsp genes. Furthermore, pre-treatment of cells with AM-2 or A. montana extract decreased melanin production and expression and activity of tyrosinase.


CONCLUSIONS
These results suggest that AM-2 and A. montana extract could be beneficial for use in hypopigmenting cosmetics as a consequence of their stimulatory effects on HSP70 expression.
Department of Drug Discovery and Development, Faculty of Pharmacy, Keio University, Tokyo 105-8512, Japan.
Keiko
K
Usui
Faculty of Pharmaceutical Sciences, Sojo University, Kumamoto 860-0082, Japan.
Tsuyoshi
T
Ikeda
Department of Drug Discovery and Development, Faculty of Pharmacy, Keio University, Tokyo 105-8512, Japan.
Yuumi
Y
Horibe
Department of Drug Discovery and Development, Faculty of Pharmacy, Keio University, Tokyo 105-8512, Japan.
Mitsuyoshi
M
Nakao
Department of Drug Discovery and Development, Faculty of Pharmacy, Keio University, Tokyo 105-8512, Japan.
Tatsuya
T
Hoshino
Department of Drug Discovery and Development, Faculty of Pharmacy, Keio University, Tokyo 105-8512, Japan. Electronic address: mizushima-th@pha.keio.ac.jp.
Tohru
T
Mizushima
D000818:Animals; D018650:Arnica / Q000737:chemistry; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D000431:Ethanol / Q000737:chemistry; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism*; D008543:Melanins / Q000096:biosynthesis; D008546:Melanoma, Experimental; D051379:Mice; D014442:Monophenol Monooxygenase / Q000378:metabolism; D008517:Phytotherapy; D010936:Plant Extracts / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D010946:Plants, Medicinal; D012867:Skin / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D062327:Skin Lightening Preparations / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D012880:Skin Pigmentation / Q000187:drug effects*; D012997:Solvents / Q000737:chemistry; D015854:Up-Regulation
D018840:HSP70 Heat-Shock Proteins; D008543:Melanins; D010936:Plant Extracts; D062327:Skin Lightening Preparations; D012997:Solvents; D000431:Ethanol; D014442:Monophenol Monooxygenase
Arnica montana; Heat shock protein 70; Hypopigmenting cosmetics; Melanogenesis; Photo-aging