Lü Suyuan, Shang Bingqing, Liu Guolong, Sun Luyan, Wu Qiu, Geng Yue
Key Laboratory of Food Nutrition and Safety of SDNU, Provincial Key Laboratory of Animal Resistant Biology, College of Life Science, Shandong Normal University, Jinan, Shandong 250358, P.R. China.
Biomed Rep. 2025 Jun 6;23(2):133. doi: 10.3892/br.2025.2011. eCollection 2025 Aug.
The present study aimed to investigate the melanin synthesis effect of pine needle essential oil (PNEO) extracted using microwave-assisted extraction on α-MSH-induced B16F10 melanoma cells and its molecular mechanism of action. Cell Counting Kit-8 was used to measure the safe concentration of B16F10 cells after 24 and 48 h post-treatment. Tyrosinase (TYR) activity assay was used to examine intracellular enzyme viability; western blotting (WB) and reverse transcription-quantitative (RT-qPCR) assays were utilized to assess the transcription and expression of genes and proteins associated with melanogenesis. Relevant targets in the cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA) signaling pathway were verified by the addition of target inhibitors or activators. Results showed that PNEO decreased the levels of melanin and the activity of TYR in B16F10 cells. RT-qPCR and WB results showed that PNEO downregulated the expression of melanogenesis-related genes and proteins such as microphthalmia-associated transcription factor, TYR, TYR-related protein-1 and melanocortin 1 receptor, and reduced the levels of phosphorylated PKA and phosphorylated cyclic-AMP response binding protein. These results suggested that the inhibitory effect of PNEO on melanin production may be related to the cAMP/PKA pathway. Verification through the addition of target inhibitors or activators confirmed that PNEO regulates melanin synthesis and TYR activity through the cAMP/PKA signaling pathway.
本研究旨在探讨微波辅助萃取法提取的松针精油（PNEO）对α-MSH诱导的B16F10黑色素瘤细胞黑色素合成的影响及其分子作用机制。采用细胞计数试剂盒-8检测处理后24小时和48小时B16F10细胞的安全浓度。采用酪氨酸酶（TYR）活性测定法检测细胞内酶活性；利用蛋白质免疫印迹法（WB）和逆转录定量（RT-qPCR）测定法评估与黑色素生成相关的基因和蛋白质的转录和表达。通过添加靶标抑制剂或激活剂验证环磷酸腺苷（cAMP）/蛋白激酶A（PKA）信号通路中的相关靶点。结果表明，PNEO可降低B16F10细胞中黑色素水平和TYR活性。RT-qPCR和WB结果显示，PNEO下调小眼相关转录因子、TYR、TYR相关蛋白-1和黑皮质素1受体等黑色素生成相关基因和蛋白质的表达，并降低磷酸化PKA和磷酸化环磷酸腺苷反应结合蛋白的水平。这些结果表明，PNEO对黑色素生成的抑制作用可能与cAMP/PKA通路有关。通过添加靶标抑制剂或激活剂进行验证，证实PNEO通过cAMP/PKA信号通路调节黑色素合成和TYR活性。
