Nok-Hyun Park

Inhibition effect of Gynura procumbens extract on UV-B-induced matrix-metalloproteinase expression in human dermal fibroblasts

ETHNOPHARMACOLOGICAL RELEVANCE Gynura procumbens Merr. (Asteraceae) has been used as a traditional remedy for various skin diseases in certain areas of Southeast Asia. AIM OF THE STUDY In order to evaluate the protective activity of Gynura procumbens extract on skin photoaging and elucidate its mode of action. MATERIALS AND METHODS Matrix-metalloproteinase (MMP)-1 and -9 expressions were induced by UV-B irradiation in human primary dermal fibroblasts. MMP-1 expression level was measured by enzyme-linked immunosorbent assay (ELISA) and Western blot analysis. Zymography was employed for evaluating the enzymatic activity of MMP-9. Anti-inflammatory activity and anti-oxidative capacity of the extract were evaluated by ELISA and dichlorodihydrofluorescein diacetate (DCF-DA) assay. RESULTS The ethanolic extract of Gynura procumbens inhibited MMP-1 expression up to 70% compare to negative control group. The enzymatic activity of MMP-9 was inhibited around 73% by the treatment of 20μg/mL of the extract. The extract markedly reduced the production of reactive oxygen species (ROS). Gynura procumbens extract showed an inhibitory effect on releasing pro-inflammatory cytokines (IL-6 and IL-8) in human HaCat keratinocyte. CONCLUSION The ethanolic extract of Gynura procumbens inhibited MMP-1 and MMP-9 expressions induced by UV-B irradiation via inhibition of pro-inflammatory cytokine mediator release and ROS production.

Amentoflavone Inhibits UVB-Induced Matrix Metalloproteinase-1 Expression Through the Modulation of AP-1 Components in Normal Human Fibroblasts

Amentoflavone is a well-known biflavonoid that has diverse biological effects. Previously, we reported that amentoflavone suppressed UVB-induced matrix metalloproteinase-1 (MMP-1) expression in normal human fibroblasts (NHF). We investigated the effects of amentoflavone on UVB-induced MMP-1 expression in order to elucidate its mode of action. NHF were treated with amentoflavone for indicated times and doses with UVB irradiation. The expressions of MMP-1 gene and protein were determined by RT-PCR and ELISA, respectively. MAP kinase phosphorylation and the expression of c-Fos protein were determined by Western blot. The treatment of amentoflavone completely blocked the upregulation of MMP-1 which is induced by UVB irradiation in HaCaT–NHF co-culture in a dose-dependent manner as well as in NHF monoculture. Also, amentoflavone inhibited UVB-induced activation of extracellular signal-regulated kinase (ERK) without changing total ERK protein level, and did not affect p38 or JNK activation. Finally, AP-1 transcription factor components, phospho-c-Jun and c-Fos protein expressions were decreased by amentoflavone treatment. The major finding of this study shows that amentoflavone inhibits intracellular cell signaling ERK pathway leading to the prevention of MMP-1 expression in human skin fibroblasts. Therefore, these results strongly suggest that amentoflavone should be investigated as a potential agent for the prevention and the treatment of skin photoaging.

Different effects of five depigmentary compounds, rhododendrol, raspberry ketone, monobenzone, rucinol and AP736 on melanogenesis and viability of human epidermal melanocytes

Numerous medications are used to treat hyperpigmentation. However, several reports have indicated that repeated application of some agents, such as rhododendrol (RD), raspberry ketone (RK) and monobenzone (MB), can be toxic to melanocytes. Although these agents had severe side effects in human trials, no current in vitro methods can predict the safety of such drugs. This study assessed the in vitro effects of five depigmentary compounds including leukoderma‐inducing agents. In particular, we determined the effects of different concentrations and exposure times of different depigmentary agents on cell viability and melanogenesis in the presence and absence of ultraviolet B (UVB) radiation. Concentrations of RD, RK and MB that inhibit melanogenesis are similar to concentrations that are cytotoxic; however, concentrations of rucinol (RC) and AP736 that inhibit melanogenesis are much lower than concentrations that are cytotoxic. Furthermore, the concentrations that cause toxic effects depend on exposure duration, and prolonged exposure to RD, RK and MB had more cytotoxic effects than prolonged exposure to RC and AP736. The cytotoxic effects of RD and RK appear to be mediated by apoptosis due to increased expression of caspase‐3 and caspase‐8; UVB radiation increased the cytotoxicity of these agents and also increased caspase activity. Our results indicate that different leukoderma‐inducing compounds have different effects on the viability of normal epidermal melanocytes and suggest that the in vitro assay used here can be used to predict whether an investigational compound that induces leukoderma may lead to adverse effects in human trials.

Alstonia scholaris R. Br. Significantly Inhibits Retinoid-Induced Skin Irritation In Vitro and In Vivo

Topical retinoids inhibit matrix metalloproteinases and accelerate collagen synthesis, thereby triggering antiaging effects in the skin. However, topical retinoids can cause severe skin reactions, including scaling, erythema, papules, and inflammation. The present study demonstrates that the ethanolic bark extract of Alstonia scholaris R. Br. can significantly inhibit all-trans retinoic acid-induced inflammation in human HaCat keratinocyte cells. Furthermore, two representative retinoid-induced proinflammatory cytokines, monocyte chemoattractant protein-1 and interleukin-8, were significantly suppressed by A. scholaris extract (by 82.1% and 26.3% at 100 ppm, and dose-dependently across the tested concentrations) in vitro. In a cumulative irritation patch test, A. scholaris extract decreased retinol-induced skin irritation, while strengthening the ability of retinoids to inhibit matrix metalloproteinase-1 expression, which is strongly associated with aging effects. These results suggest that A. scholaris is a promising compound that may increase the antiaging function of retinoids while reducing their ability to cause skin irritation.

Activation of transient receptor potential melastatin 8 reduces ultraviolet B-induced prostaglandin E2 production in keratinocytes

Transient receptor potential melastatin 8 (TRPM8) is a member of the TRP family, and is activated at temperatures below 22°C, or by cooling compounds such as menthol. In this study, it was found that a new role of TRPM8 activation on prostaglandin E2 (PGE2), an inflammatory cytokine and dendritogenesis stimulator of normal human melanocytes. Normal human keratinocytes were pretreated with menthol or incubated at 22°C for TRPM8 activation before ultraviolet (UV)‐B irradiation. To examine the specificity between TRPM8 activation and PGE2 release, we inhibited TRPM8 with the antagonist (capsazepine), or introduced TRPM8 siRNA for a gene silencing experiment. UV‐B irradiation significantly induced PGE2 release in normal human keratinocytes. Interestingly, activation of TRPM8 at 22°C or with menthol inhibited UV‐B‐induced PGE2 release. The effect of the TRPM8 agonist was completely blocked by pretreatment with the TRPM8 antagonist, capsazepine. When TRPM8 expression was suppressed by siRNA, UV‐B irradiation still upregulated PGE2 in keratinocytes, but pretreatment of menthol or low temperature did not inhibit UV‐B‐induced PGE2. In conclusion, the activation of TRPM8 inhibits UV‐B‐induced PGE2 production in keratinocytes, and the activation of TRPM8 may reduce inflammatory responses in skin.

Antimelanogenic Efficacy of Melasolv (3,4,5-Trimethoxycinnamate Thymol Ester) in Melanocytes and Three-Dimensional Human Skin Equivalent

Background/Aims: Excessive melanogenesis often causes unaesthetic hyperpigmentation. In a previous report, our group introduced a newly synthesized depigmentary agent, Melasolv™ (3,4,5-trimethoxycinnamate thymol ester). In this study, we demonstrated the significant whitening efficacy of Melasolv using various melanocytes and human skin equivalents as in vitro experimental systems. Methods: The depigmentary effect of Melasolv was tested in melan-a cells (immortalized normal murine melanocytes), α-melanocyte-stimulating hormone (α-MSH)-stimulated B16 murine melanoma cells, primary normal human melanocytes (NHMs), and human skin equivalent (MelanoDerm). The whitening efficacy of Melasolv was further demonstrated by photography, time-lapse microscopy, Fontana-Masson (F&M) staining, and 2-photon microscopy. Results: Melasolv significantly inhibited melanogenesis in the melan-a and α-MSH-stimulated B16 cells. In human systems, Melasolv also clearly showed a whitening effect in NHMs and human skin equivalent, reflecting a decrease in melanin content. F&M staining and 2-photon microscopy revealed that Melasolv suppressed melanin transfer into multiple epidermal layers from melanocytes as well as melanin synthesis in human skin equivalent. Conclusion: Our study showed that Melasolv clearly exerts a whitening effect on various melanocytes and human skin equivalent. These results suggest the possibility that Melasolv can be used as a depigmentary agent to treat pigmentary disorders as well as an active ingredient in cosmetics to increase whitening efficacy.

Metabolomics approach for understanding geographical dependence of soybean leaf metabolome

The soybean plant (Glycine max) is widely used as an ingredient in various foods, nutraceuticals and cosmetics, due to their diverse bioactive compounds. Their metabolic compositions are likely affected by environmental conditions during growth. To investigate the influence of different environmental conditions on the metabolite composition of soybean leaves, we cultivated soybean (G. max Sinhwa) in the southernmost island and volcanic region of Korea, and in the central section and limestone region of the Korean peninsula. Comprehensive metabolite variations of their leaves were analyzed through 1H NMR-based metabolomics approach. With marked differences in soil compositions and climatic conditions between the two growing areas, differences in accumulations of pinitol and diverse flavonoids were noted between the soybean leaves, reflecting the distinct metabolism of soybean plants for physiological adaptation toward different environmental conditions. Therefore, the current study highlights the geographical dependences of diverse soybean leaf metabolites for developing biofunction-enhanced soybean products.