Leathesia difformis Extract Inhibits α-MSH-Induced Melanogenesis in B16F10 Cells via Down-Regulation of CREB Signaling Pathway.

Leathesia difformis (L.) Areschoug (L. difformis) is a species of littoral brown algae of the class Phaeophyceae. Only a few studies on the apoptotic, antiviral, and antioxidant properties of L. difformis have been reported, and its inhibitory effect on melanin synthesis has not been studied. The aim of this study was to investigate the anti-melanogenic effect of L. difformis extract on α-melanocyte-stimulating hormone (α-MSH)-induced B16F10 melanocytes and its mechanism of action. L. difformis was extracted using 80% ethanol (LDE) and then fractioned between ethyl acetate (LDE-EA) and water (LDE-A). Our data demonstrated that LDE-EA significantly inhibited melanin level and cellular tyrosinase activity in α-MSH-stimulated B16 cells. In addition, the expression of genes associated with melanin synthesis, such as microphthalmia-associated transcription factor (Mitf), tyrosinase (Tyr), tyrosinase-related protein-1 (Trp-1), dopachrome tautomerase (Dct), and melanocortin 1 receptor (Mc1r) was down-regulated by LDE-EA treatment. Moreover, LDE-EA decreased p-CREB levels, which suggests that the inhibition of the cAMP/PKA/CREB pathways may be involved in the anti-melanogenic effect of LDE-EA. Thus, this study revealed that LDE-EA is an effective inhibitor of hyperpigmentation through inhibition of CREB pathways and may be considered as a potential therapeutic agent for hyperpigmentation disorders.

A Novel Synthetic Material, BMM, Accelerates Wound Repair by Stimulating Re-Epithelialization and Fibroblast Activation.

Cutaneous wound repair is an intricate process whereby the skin reprograms itself after injury. In the mid-phase of wound repair, the proliferation, migration, and differentiation of cells are the major mechanisms to lead remodeling. We investigated the effect of BMM ((1E,2E)-1,2-bis((6-bromo-2H-chromen-3-yl)methylene)hydrazine), a novel synthetic material, on the migration and viability of keratinocytes or fibroblasts using the in vitro scratch woundhealing, electric cell-substrate imedance sensing (ECIS), invasion, and MTT assays. Cell migration-related factors were analyzed using western blot, and we found that treatment with BMM stimulated the EMT pathway and focal adhesion kinase (FAK)/Src signaling. Differentiation of HaCaT keratinocyte and fibroblast cells was also stimulated by BMM and specifically, NOX2/4 contributed to the activation of fibroblasts for wound healing. Furthermore, BMM treated HaCaT keratinocyte and fibroblast-co-cultured cells increased migration and differentiation. TGF-ß and Cyr61 were also secreted to a greater extent than in single cultured cells. In vivo experiments showed that treatment with BMM promotes wound closure by promoting re-epithelialization. In this study, we demonstrated that a novel synthetic material, BMM, is capable of promoting wound healing via the stimulation of re-epithelialization in the epidermis and the activation of fibroblasts in the dermis, in particular, via the acceleration of the interaction between the epidermis and dermis.

TMF and glycitin act synergistically on keratinocytes and fibroblasts to promote wound healing and anti-scarring activity.

Keratinocyte-fibroblast interactions are critical for skin repair after injury. During the proliferative phase of wound healing, proliferation, migration and differentiation of these cells are the major mechanisms leading to tissue remodeling. We have previously reported that glycitin, a major soy isoflavone, stimulates dermal fibroblast proliferation; and the phytochemical, 4',6,7-trimethoxyisoflavone (TMF), induces migration of HaCaT keratinocyte cells. We therefore investigated whether these compounds display synergistic effects on skin cells during wound healing in vitro and in vivo. Co-treatment with TMF and glycitin synergistically promotes the proliferation and migration of both keratinocytes and dermal fibroblasts, with a 1:1 ratio of these compounds showing the greatest efficacy in our co-culture system. This keratinocyte-fibroblast interaction occurred via the secretion of TGF-ß, and the induction of differentiation and proliferation was confirmed in both indirect and direct co-culture assays. In an excisional and burn wound animal model, mice treated with a 1:1 ratio of TMF and glycitin showed faster wound closure, regeneration and scar reduction than even the positive control drug. These data indicate that two isoflavones, TMF and glycitin, act synergistically to promote wound healing and anti-scarring and could potentially be developed together as a bioactive therapeutic for wound treatment.

A synthetic isoflavone, DCMF, promotes human keratinocyte migration by activating Src/FAK signaling pathway.

Flavonoids are plant secondary compounds with various pharmacological properties. We previously showed that one flavonoid, trimethoxyisoflavone (TMF), could promote wound healing by inducing keratinocyte migration. Here, we screened TMF derivatives for enhanced activity and identified one compound, 2',6 Dichloro-7-methoxyisoflavone (DCMF), as most effective at promoting migration in a scratch wound assay. Using the HaCaT keratinocyte cell line, we found DCMF treatment induced phosphorylation of both FAK and Src, and increased keratinocyte migration. DCMF-induced Src kinase could promote activation of ERK, AKT, and p38 signaling pathways, and DCMF-induced secretion of matrix metalloproteinase (MMP)-2 and MMP-9 and partial epithelial-mesenchymal transition (EMT), whereas Src inhibition abolished DCMF-induced EMT. Using an in vivo excisional wound model, we observed improved wound closure and re-epithelialization in DCMF-treated mice, as compared to controls. Collectively, our data demonstrate that DCMF induces cell migration and promotes wound healing through activation of Src/FAK, ERK, AKT, and p38 MAPK signaling.

Novel naphthochalcone derivative accelerate dermal wound healing through induction of epithelial-mesenchymal transition of keratinocyte.

BACKGROUND: Wound healing is an intricate process whereby the skin repairs itself after injury. The epithelial-mesenchymal transition (EMT) is associated with wound healing and tissue regeneration. Naphthochalcone derivatives have various pharmaceutical properties. We investigated the effect of a novel naphthochalcone derivative, 2-(5-(2,4,6-trimethoxyphenyl)-4,5-dihydro-1H-pyrazol-3-yl)naphthalen-1-ol (TDPN), on dermal wound healing in vivo and the migration of keratinocytes in vitro. RESULT: We investigated the effect of TDPN on signaling pathway and epithelial-mesenchymal transition through protein and transcriptional expression. The TDPN treatment accelerated dermal closure about 3 days and remodeling of dermis. We found that treatment with TDPN induced the migration of keratinocytes but not cytotoxicity. TDPN induced the phosphorylation of ERK and AKT. TDPN-treated cells showed loss of adherence protein and showed induction of the transcriptional factor Slug, mesenchymal marker, and fibronectin. Moreover, TDPN treatment induced the expression of matrix metalloproteinase-1 (MMP-1), which degrades specific components of the extracellular matrix, thereby providing new substrates that facilitate migration and invasion. MMP expression is considered to be one of the major attributes acquired by cells after EMT. CONCLUSION: We propose that a novel naphthochalcone derivative TDPN is capable of promoting keratinocyte migration via the induction of EMT resulting acceleration of wound closure and matrix remodeling.

Winogradskyella eckloniae sp. nov., a marine bacterium isolated from the brown alga Ecklonia cava.

A novel bacterial strain, designated EC29(T), was isolated from the brown alga Ecklonia cava collected on Jeju Island, Republic of Korea. Cells of strain EC29(T) were Gram-stain-negative, aerobic, rod-shaped and motile by gliding. Growth was observed at 10-30 °C (optimum, 20-25 °C), at pH 6.0-9.5 (optimum, pH 7.5) and in the presence of 1-5% (w/v) NaCl. Phylogenetic analyses based on the 16S rRNA gene sequence revealed that the strain belonged to the genus Winogradskyella. Strain EC29(T) exhibited the highest 16S rRNA gene sequence similarities, of 96.5-97.8%, to the type strains of Winogradskyella pulchriflava EM106(T), Winogradskyella echinorum KMM 6211(T) and Winogradskyella ulvae KMM 6390(T). Strain EC29(T) exhibited < 27% DNA-DNA relatedness with Winogradskyella pulchriflava EM106(T) and Winogradskyella echinorum KMM 6211(T). The predominant fatty acids of strain EC29(T) were iso-C15 : 0, iso-C15 : 1 G, C15 : 0, iso-C17 : 0 3-OH, iso-C15 : 0 3-OH and anteiso-C15 : 0. The DNA G+C content was 31.1 mol% and the major respiratory quinone was menaquinone-6 (MK-6). Based on a polyphasic study, strain EC29(T) is considered to represent a novel species of the genus Winogradskyella, for which the name Winogradskyella eckloniae sp. nov. is proposed. The type strain is EC29(T) ( = KCTC 32172(T) = JCM 18703(T)).

Positive conversion of tuberculin skin test and performance of interferon release assay to detect hidden tuberculosis infection during anti-tumor necrosis factor agent trial.

OBJECTIVES: To evaluate tuberculin skin tests (TST) and interferon-gamma (IFN-gamma) assay in the detection of latent tuberculosis (TB) infection during tumor necrosis factor (TNF) antagonist treatment in Korean patients with initial negative TST result. METHODS: Eighty-six patients with rheumatic diseases who had received anti-TNF agents for over one year were investigated. Clinical data were obtained from medical records. All patients received followup TST, and IFN-gamma assay was performed in 64. RESULTS: The study population consisted of 40 rheumatoid arthritis (RA), 34 ankylosing spondylitis (AS), 9 juvenile rheumatoid arthritis (JRA), and 3 other patients. The TST converted to positive in 28 (32.6%) patients. There was no significant variation between TST conversion rate and all risk factors. Although there was no statistical significance, the odds of the TST conversion rate tended to increase with the duration of TNF antagonist administration. Nine (14.1%) of 64 patients who performed an IFN-gamma assay had positive results. Among 28 TST positive conversion cases, 4 patients with AS and 1 with psoriatic arthritis had positive IFN-gamma assay results, and one of them developed miliary TB. However, none of the 4 RA patients with positive IFN-gamma assay showed TST conversion. There was 68.6% agreement (kappa = 0.29, p = 0.02) between TST and IFN-gamma assay results. CONCLUSION: Serial TST with IFN-gamma assay may be useful to identify false-negative response to cases of latent Mycobacterium tuberculosis infection and new TB infections in patients with immune mediated inflammatory diseases during longterm anti-TNF therapy, especially in areas with intermediate TB burden.