Kumamoto-shi, Japan
Kumamoto-shi, Japan

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Yamashita Y.,Kumamoto University | Hoshino T.,Kumamoto University | Matsuda M.,Kumamoto University | Kobayashi C.,Kumamoto University | And 9 more authors.
Experimental Dermatology | Year: 2010

Skin hyperpigmentation disorders as a result of abnormal melanin production induced by ultraviolet (UV) irradiation are both a clinical and a cosmetic problem. This melanin production is mediated by tyrosinase whose expression is positively regulated by microphthalmia-associated transcription factor (MITF). We recently found that expression of heat shock protein 70 (HSP70) inhibits melanin production. In this study, we searched for HSP70 inducers from Chinese herbs and selected an ethanol extract of Eupatorium lindleyanum (E. lindleyanum). Not only melanin production but also the activity and expression of tyrosinase were significantly suppressed in cells treated with E. lindleyanum extract as well as in HSP70-overexpressing cells. The expression of MITF was clearly suppressed in cells treated with E. lindleyanum extract but not in HSP70-overexpressing cells. These results suggest that E. lindleyanum extract suppresses the expression of tyrosinase and melanin production through both HSP70-dependent and HSP70-independent mechanisms. © 2010 John Wiley & Sons A/S.


Yamashita Y.,Keio University | Yamashita Y.,Kumamoto University | Ikeda T.,Sojo University | Matsuda M.,Kumamoto University | And 5 more authors.
Biochemical Pharmacology | Year: 2012

The expression of heat shock proteins (HSPs), particularly HSP70, provides resistance to stressors. We recently reported that ultraviolet (UV)-induced melanin production and skin damage were suppressed in transgenic mice expressing HSP70 and that an extract of Eupatorium lindleyanum induces the expression of HSP70 in cells. Here we report the purification of eupalinolide A and B (EA and EB) from E. lindleyanum, and describe their actions as HSP-inducers. EA and EB both induced the expression of HSP70 in cells at concentrations that did not significantly affect cell viability. Treatment of cells with EA or EB activated heat shock factor 1 (HSF1), while the artificial suppression of HSF1 expression diminished the EA- or EB-mediated induction of HSP70 expression. Furthermore, EB inhibited the interaction between HSF1 and HSP90, which is known to inhibit the activity of HSF1. These findings suggest that EA and EB induce the expression of HSP70 via the activation of HSF1 by inhibiting the interaction between HSF1 and HSP90. EA and EB both induced the expression of HSP70 synergistically with other stressors. Furthermore, pre-treatment of cells with EA or EB suppressed melanin production and stressor-induced apoptosis. These effects were suppressed by the artificial suppression of HSP70 expression. In vivo, the percutaneous administration of EB induced the expression of HSP70 and suppressed UVB radiation-induced damage, inflammatory responses and melanin production in the skin. These results suggest that EA and EB could be beneficial for use in cosmetics and medicines as a consequence of their inhibitory action on UV-induced skin damage and melanin production. © 2012 Elsevier Inc. All rights reserved.


Matsuda M.,Kumamoto University | Hoshino T.,Kumamoto University | Yamashita Y.,Kumamoto University | Tanaka K.-I.,Kumamoto University | And 7 more authors.
Journal of Biological Chemistry | Year: 2010

Irradiation with UV light, especially UVB, causes epidermal damage via the induction of apoptosis, inflammatory responses, and DNA damage. Various stressors, including UV light, induce heat shock proteins (HSPs) and the induction, particularly that of HSP70, provides cellular resistance to such stressors. The anti-inflammatory activity of HSP70, such as its inhibition of nuclear factor kappa B (NF-κB), was recently revealed. These in vitro results suggest that HSP70 protects against UVB-induced epidermal damage. Here we tested this idea by using transgenic mice expressing HSP70 and cultured keratinocytes. Irradiation of wild-type mice with UVB caused epidermal damage such as induction of apoptosis, which was suppressed in transgenic mice expressing HSP70. UVB-induced apoptosis in cultured keratinocytes was suppressed by overexpression of HSP70. Irradiation of wild-type mice with UVB decreased the cutaneous level of IκB-α (an inhibitor of NF-κB) and increased the infiltration of leukocytes and levels of pro-inflammatory cytokines and chemokines in the epidermis. These inflammatory responses were suppressed in transgenic mice expressing HSP70. In vitro, the overexpression of HSP70 suppressed the expression of pro-inflammatory cytokines and chemokines and increased the level of IκB-α in keratinocytes irradiated with UVB. UVB induced an increase in cutaneous levels of cyclobutane pyrimidine dimers and 8-hydroxy-2′-deoxyguanosine, both of which were suppressed in transgenic mice expressing HSP70. This study provides genetic evidence that HSP70 protects the epidermis from UVB-induced radiation damage. The findings here also suggest that the protective action of HSP70 is mediated by anti-apoptotic, anti-inflammatory, and anti-DNA damage effects. © 2010 by The American Society for Biochemistry and Molecular Biology, Inc.


Hoshino T.,Kumamoto University | Matsuda M.,Kumamoto University | Yamashita Y.,Kumamoto University | Takehara M.,Kumamoto University | And 8 more authors.
Journal of Biological Chemistry | Year: 2010

Skin hyperpigmentation disorders due to abnormal melanin production induced by ultraviolet (UV) irradiation are both a clinical and cosmetic problem. UV irradiation stimulates melanin production in melanocytes by increasing intracellular cAMP. Expression of heat shock proteins (HSPs), especially HSP70, is induced by various stressors, including UV irradiation, to provide cellular resistance to such stressors. In this study we examined the effect of expression of HSP70 on melanin production both in vitro and in vivo. 3-Isobutyl-1- methylxanthine (IBMX), a cAMP-elevating agent, stimulated melanin production in cultured mouse melanoma cells, and this stimulation was suppressed in cells overexpressing HSP70. IBMX-dependent transcriptional activation of the tyrosinase gene was also suppressed in HSP70-overexpressing cells. Expression of microphthalmia-associated transcription factor (MITF), which positively regulates transcription of the tyrosinase gene, was up-regulated by IBMX; however, this up-regulation was not suppressed in HSP70-overexpressing cells. On the other hand, immunoprecipitation and immunostaining analyses revealed a physical interaction between and co-localization of MITF and HSP70, respectively. Furthermore, the transcription of tyrosinase gene in nuclear extract was inhibited by HSP70. In vivo, UV irradiation of wild-type mice increased the amount of melanin in the basal layer of the epidermis, and this increase was suppressed in transgenic mice expressing HSP70. This study provides the first evidence of an inhibitory effect of HSP70 on melanin production both in vitro and in vivo. This effect seems to be mediated by modulation of MITF activity through a direct interaction between HSP70 and MITF. © 2010 by The American Society for Biochemistry and Molecular Biology, Inc.

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