School of Bioscience and Biotechnology, Tokyo University of Technology, Hachioji, Japan.
Arch Dermatol Res. 2012 Dec;304(10):803-16. doi: 10.1007/s00403-012-1248-y. Epub 2012 May 26.
We established a model for the stem cell factor (SCF)-associated stimulation of human epidermal equivalent (HEE) pigmentation. The addition of SCF (at 5 nM) gradually stimulated the visible pigmentation of HEEs over 14 days of treatment. A time course study using real-time RT-PCR and western blotting analysis demonstrated that the expression of all melanocyte-specific genes and proteins examined was gradually up-regulated over 7-10 days of treatment with SCF. The addition of astaxanthin (Ax) at concentrations of 1, 4, or 8 μM markedly abolished the SCF- but not the endothelin (EDN)1-elicited increase in visible pigmentation over 14 days in a dose-dependent manner, with almost complete inhibition at 8 μM. While no degeneration of the epidermal tissue was visible at day 14 by HE staining, melanin deposition throughout the epidermis was markedly reduced in the Ax-treated HEEs at day 14 compared to untreated controls. Ax significantly reduced the eumelanin content of HEEs to the non-SCF-stimulated level at concentrations of 4 or 8 μM compared with untreated controls. Real-time RT-PCR and western blotting of Ax-treated HEEs revealed that the SCF-stimulated expression of tyrosinase (TYR), TYR-related protein-1 (TYRP1), and Pmel17, as well as microphthalmia-associated transcription factor (MITF), is significantly suppressed by Ax at the transcriptional and translational levels. Studies using cultured normal human melanocytes revealed that pre-treatment with Ax interrupts the SCF- but not the EDN1-induced stimulation of TYR activity, and there was no direct inhibitory effect of Ax on TYR activity in vitro. These findings indicate that Ax attenuates SCF-stimulated pigmentation by directly interrupting SCF-associated intracellular signaling linkages through increased expression of MITF, which leads to the stimulated expression of melanogenic genes and proteins in a reactive oxygen species depletion-independent mechanism.
我们建立了一个模型，用于研究干细胞因子（SCF）对人表皮等效物（HEE）色素沉着的刺激作用。添加 SCF（5 nM）逐渐刺激 HEE 在 14 天的治疗过程中可见的色素沉着。使用实时 RT-PCR 和 Western blot 分析的时间过程研究表明，在 SCF 处理 7-10 天后，所有检查的黑素细胞特异性基因和蛋白质的表达逐渐上调。虾青素（Ax）的添加浓度为 1、4 或 8 μM，以剂量依赖性方式显著消除了 SCF-但不是内皮素（EDN1）引发的可见色素沉着增加，在 8 μM 时几乎完全抑制。通过 HE 染色，在第 14 天未见表皮组织退化，但与未处理的对照相比，Ax 处理的 HEE 中整个表皮的黑色素沉积明显减少。与未处理的对照相比，Ax 在 4 或 8 μM 浓度下将 HEE 的真黑素含量显著降低至非 SCF 刺激水平。Ax 处理的 HEE 的实时 RT-PCR 和 Western blot 显示，SCF 刺激的酪氨酸酶（TYR）、TYR 相关蛋白-1（TYRP1）和 Pmel17 以及小眼畸形相关转录因子（MITF）的表达在转录和翻译水平上均被 Ax 显著抑制。使用培养的正常人黑素细胞进行的研究表明，Ax 的预处理可中断 SCF-但不中断 EDN1 诱导的 TYR 活性刺激，并且 Ax 对 TYR 活性没有直接的抑制作用在体外。这些发现表明，Ax 通过增加 MITF 的表达，直接中断与 SCF 相关的细胞内信号通路，从而减弱 SCF 刺激的色素沉着，这导致在活性氧耗竭非依赖性机制中刺激黑素生成基因和蛋白质的表达。
