Grabacka Maja, Wieczorek Justyna, Michalczyk-Wetula Dominika, Malinowski Michel, Wolan Norbert, Wojcik Katarzyna, Plonka Przemyslaw M
Department of Food Biotechnology, Faculty of Food Technology, University of Agriculture, ul. Balicka 122, 30-149, Kraków, Poland.
Department of Biophysics, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, ul. Gronostajowa 7, 30-387, Kraków, Poland.
Arch Dermatol Res. 2017 Apr;309(3):141-157. doi: 10.1007/s00403-016-1711-2. Epub 2017 Jan 13.
Recent studies revealed the cooperation between peroxisome proliferator-activated receptor gamma (PPARγ) and α-MSH signaling, which results in enhanced melanogenesis in melanocytes and melanoma cells. However, the agonists of PPARα, such as fenofibrate, exert depigmenting effect. Therefore, we aimed to check how the PPARα expression level affects the antimelanogenic activity of fenofibrate and whether PPARα modulates melanogenesis independently of its agonist. To answer these questions, we used three B16 F10-derived cell lines, which varied in the PPARα expression level and were developed by stable transfection with plasmids driving shRNA-based PPARα silencing or overexpression of PPARα-emerald GFP fusion protein. Melanin contents were assessed with electron paramagnetic resonance spectroscopy along with color component image analysis-a novel approach to pigment content characteristics in melanoma cells. B16 F10 wt and Ctrl shRNA lines showed intermediate pigmentation, whereas the pigmentation of the B16 F10-derived cell lines was inversely correlated with the PPARα expression level. We observed that cells overexpressing PPARα were almost amelanotic and cells with reduced PPARα protein level were heavily melanized. Furthermore, fenofibrate down-regulated the melanogenic apparatus (MITF, tyrosinase, and tyrosinase-related proteins) in the cells with the regular PPARα expression level resulting in their visibly lower total melanin content in all the cell lines. From these observations, we conclude that fenofibrate works as a strong depigmenting agent, which acts independently of PPARα, but in an additive fashion. Our results also indicate that alterations in PGC-1a acetylation and expression level might contribute to the regulation of melanogenesis by PPARα and fenofibrate.
最近的研究揭示了过氧化物酶体增殖物激活受体γ（PPARγ）与α-促黑素（α-MSH）信号之间的协同作用，这会导致黑素细胞和黑色素瘤细胞中的黑素生成增强。然而，PPARα的激动剂，如非诺贝特，具有色素脱失作用。因此，我们旨在研究PPARα的表达水平如何影响非诺贝特的抗黑素生成活性，以及PPARα是否独立于其激动剂调节黑素生成。为了回答这些问题，我们使用了三种源自B16 F10的细胞系，它们的PPARα表达水平各不相同，是通过用驱动基于短发夹RNA（shRNA）的PPARα沉默或PPARα-翡翠绿色荧光蛋白融合蛋白过表达的质粒进行稳定转染而构建的。使用电子顺磁共振光谱以及颜色成分图像分析（一种用于黑色素瘤细胞色素含量特征分析的新方法）评估黑色素含量。B16 F10野生型和对照shRNA细胞系表现出中等色素沉着，而源自B16 F10的细胞系的色素沉着与PPARα表达水平呈负相关。我们观察到，过表达PPARα的细胞几乎无黑色素生成，而PPARα蛋白水平降低的细胞则黑色素沉着严重。此外，非诺贝特下调了具有正常PPARα表达水平的细胞中的黑素生成相关蛋白（小眼畸形相关转录因子（MITF）、酪氨酸酶和酪氨酸酶相关蛋白），导致所有细胞系中它们的总黑色素含量明显降低。从这些观察结果中，我们得出结论，非诺贝特作为一种强效的色素脱失剂，其作用独立于PPARα，但具有累加效应。我们的结果还表明，过氧化物酶体增殖物激活受体γ共激活因子1α（PGC-1α）乙酰化和表达水平的改变可能有助于PPARα和非诺贝特对黑素生成的调节。
