School of Bioscience and Biotechnology, Tokyo University of Technology, Tokyo, Japan.
Phytother Res. 2011 Sep;25(9):1398-411. doi: 10.1002/ptr.3552. Epub 2011 Jun 16.
Redox imbalances have been shown to be closely linked to a variety of altered cellular responses and profoundly affect intracellular signaling pathways, especially the PKC/MAPK pathway which is a major pathway involved in regulating melanogenesis within human melanocytes. To elucidate the effects of redox balance regulation on epidermal hyperpigmentary disorders, an antioxidant-rich herb extract of Withania somnifera was used to assess its effect on endothelin-1 (EDN1)-stimulated pigmentation in human epidermis equivalents and its biological mechanisms analysed. Addition of the Withania somnifera extract (10 µg/mL) elicited a marked depigmenting effect on EDN1 (10 nm)-stimulated pigmentation which was accompanied by a significant decrease in eumelanin content. Real-time RT-PCR and western blotting revealed that the stimulated expression of melanocyte-specific mRNAs and proteins, including microphthalmia associated transcription factor (MITF), was significantly suppressed at days 7-10 of culture by the Withania somnifera extract (10 µg/mL), suggesting an impairment in intracellular signaling upstream of gene expression. Signaling analysis revealed that in Withania somnifera extract (10 µg/mL)-treated human melanoma cells in culture, there was a marked deficiency in EDN1 (10 nm)-stimulated phosphorylation of Raf-1, MEK, ERK, MITF and Cyclic AMP responsive element binding protein (CREB) at 15 min after EDN1 treatment. Consistently, treatment with withaferin A, a major component of the Withania somnifera extract, at concentrations of 10-50 µm also significantly down-regulated the EDN1 stimulated phosphorylation of Raf-1, MEK, ERK, MITF and CREB at 15 min after EDN1 treatment. Since Raf-1 is phosphorylated by protein kinase C (PKC) activity, these findings indicate that the Withania somnifera extract attenuates EDN1-stimulated pigmentation by preferentially inhibiting EDN1-triggered PKC activity.
氧化还原失衡已被证明与多种细胞反应改变密切相关，并深刻影响细胞内信号通路，特别是蛋白激酶 C/丝裂原活化蛋白激酶（PKC/MAPK）通路，该通路是调节人黑素细胞中黑色素生成的主要通路之一。为阐明氧化还原平衡调节对表皮色素沉着过度疾病的影响，使用富含抗氧化剂的茄属植物提取物来评估其对人表皮模型中内皮素-1（EDN1）刺激的色素沉着的影响，并分析其生物学机制。添加茄属植物提取物（10μg/ml）可显著抑制 EDN1（10nm）刺激的色素沉着，同时黑色素含量明显降低。实时 RT-PCR 和 Western blot 显示，在培养的第 7-10 天，茄属植物提取物（10μg/ml）显著抑制黑素细胞特异性 mRNA 和蛋白的刺激表达，包括小眼畸形相关转录因子（MITF），提示基因表达上游的细胞内信号受损。信号分析显示，在培养的人黑素瘤细胞中，茄属植物提取物（10μg/ml）处理后，EDN1（10nm）刺激的 Raf-1、MEK、ERK、MITF 和环磷酸腺苷反应元件结合蛋白（CREB）磷酸化在 EDN1 处理后 15 分钟时明显不足。一致地，用茄属植物提取物的主要成分胡椒酚 A（浓度为 10-50μm）处理也显著下调了 EDN1 刺激的 Raf-1、MEK、ERK、MITF 和 CREB 的磷酸化，EDN1 处理后 15 分钟。由于 Raf-1 被蛋白激酶 C（PKC）活性磷酸化，这些发现表明茄属植物提取物通过优先抑制 EDN1 触发的 PKC 活性来减轻 EDN1 刺激的色素沉着。
