Wang Tian-Jing, An Jing, Chen Xiao-Hui, Deng Qiu-Di, Yang Liu
TCM-Integrated Hospital, Southern Medical University, Guangzhou 510310, China; School of Traditional Chinese Medicine, Southern Medical University, Guangzhou 510515, China.
Cancer Research Institute, Southern Medical University, Guangzhou 510515, China.
J Ethnopharmacol. 2014 May 28;154(1):240-8. doi: 10.1016/j.jep.2014.04.016. Epub 2014 Apr 16.
Cuscuta chinensis seeds have traditionally been used to treat freckles and melasma in Asia, although recent reports have revealed that Semen cuscutae is a promoter of melanogenesis. The present study aims to investigate the mechanism of this opposite effect of Semen cuscutae on melanogenesis.
In accordance with traditional usage, the water fraction and the ethanol fraction from Semen cuscutae (WFSC/EFSC) were extracted to determine the herbal effects by examining the activity of mushroom tyrosinase, cellular melanin contents, tyrosinase activity assay, quantitative-reverse transcription polymerase chain reaction (qRT-PCR), and Western blot analysis for tyrosinase in B16F10 mouse melanoma cells. The melanocyte phenotypes of zebrafish larvae were observed while the in vivo melanin contents and tyrosinase activity were determined.
The activity of mushroom tyrosinase assay shown that WFSC was an uncompetitive inhibitor of mushroom tyrosinase, while EFSC indicated dose-dependent activation of the mushroom tyrosinase activity. The WFSC markedly inhibited 3-isobutyl-1-methylxanthine (IBMX)-stimulated melanin synthesis and tyrosinase activity in vitro. Howeveran accelerant role in melanin synthesis and tyosinase activity. Neither fraction had any effect on the IBMX-induced expression of tyrosinase protein or mRNA. The WFSC strongly inhibited melanin synthesis and cellular tyrosinase activity in vivo. Furthermore, with the function of WFSC at a higher concentration, a punctate melanocyte pattern appeared that was similar to the pattern induced by arbutin or Mequinol (MQ). The EFSC had no effect on the melanocytes of zebrafish larvae. It was discovered that WFSC did not show a stable inhibitory effect until it was extracted 1 month later.
These results suggest that the opposite effects of Cuscuta chinensis seeds were caused by the extraction methods and that time has an important role on the effect of WFSC. Both WFSC and EFSC significantly influence melanogenesis by regulating enzymatic activity of tyrosinase. In addition, the data indicate that wildtype (WT) zebrafish may be an ideal model for testing inhibitors of melanogenesis from clinically active herbs.
在亚洲，菟丝子种子传统上用于治疗雀斑和黄褐斑，尽管最近的报告显示菟丝子是黑色素生成的促进剂。本研究旨在探讨菟丝子对黑色素生成这种相反作用的机制。
按照传统用法，提取菟丝子的水提取物和乙醇提取物（WFSC/EFSC），通过检测蘑菇酪氨酸酶活性、细胞黑色素含量、酪氨酸酶活性测定、定量逆转录聚合酶链反应（qRT-PCR）以及对B16F10小鼠黑色素瘤细胞中酪氨酸酶进行蛋白质印迹分析来确定其药理作用。观察斑马鱼幼体的黑素细胞表型，同时测定体内黑色素含量和酪氨酸酶活性。
蘑菇酪氨酸酶活性测定表明，WFSC是蘑菇酪氨酸酶的非竞争性抑制剂，而EFSC则显示出对蘑菇酪氨酸酶活性的剂量依赖性激活。WFSC在体外显著抑制3-异丁基-1-甲基黄嘌呤（IBMX）刺激的黑色素合成和酪氨酸酶活性。然而，EFSC对黑色素合成和酪氨酸酶活性有促进作用。两种提取物对IBMX诱导的酪氨酸酶蛋白或mRNA表达均无影响。WFSC在体内强烈抑制黑色素合成和细胞酪氨酸酶活性。此外，在较高浓度的WFSC作用下，出现了点状黑素细胞模式，类似于熊果苷或甲氧基喹啉（MQ）诱导的模式。EFSC对斑马鱼幼体的黑素细胞没有影响。发现WFSC直到提取1个月后才显示出稳定的抑制作用。
这些结果表明，菟丝子种子的相反作用是由提取方法引起的，并且时间对WFSC的作用有重要影响。WFSC和EFSC均通过调节酪氨酸酶的酶活性显著影响黑色素生成。此外，数据表明野生型（WT）斑马鱼可能是测试临床活性草药中黑色素生成抑制剂的理想模型。
