七种中药乙醇提取物及补骨脂素对人黑素瘤YUGEN8细胞酪氨酸酶的影响

Effects of alcoholic extracts of seven traditional Chinese medicines and psoralen on tyrosinase in human YUGEN8 melanoma cell

目的探讨中药乙醇提取物对酪氨酸酶翻译后加工成熟及运输的影响。方法体外传代培养人无色素黑素瘤细胞YUGEN8,分别加入川芎等7味中药提取液及补骨脂素,利用蛋白免疫印迹、内切糖苷酶酶切及激光共聚焦显微镜亚细胞定位等方法,观察中药处理组细胞内酪氨酸酶的表达、成熟及内质网输出过程与阴性对照组的差异。结果与阴性对照组比,中药菟丝子及桃仁处理组细胞酪氨酸酶蛋白相对分子质量80 000左右成熟片段表达增加,能够耐受内切糖苷酶酶切；共聚焦显微镜下,酪氨酸酶的分布超出了内质网固有蛋白—钙联接蛋白的标记范围。结论中药菟丝子及桃仁对黑素瘤细胞内酪氨酸酶的成熟、稳定及内质网输出具有一定促进作用。

Objective To explore the effects of alcoholic extracts of traditional Chinese medicines on the post-translational processing and trafficking of tyrosinase. Methods Human YUGEN8 amelanotic melanoma cells were grown in vitro; the cells were incubated with one of the seven traditional Chinese medicines, including Rhizoma Chuanxiong and psoralen. Protein analysis with Western blot, enzymolysis with endoglycosidase H （ Endo H ）, and subcellular localization with laser confocal microscopy were performed. The expression, maturity and export from endoplasmic reticulum （ ER ） of tyrosinase in the treated cells were compared with those in the untreated controls. Results Compared with controls, an approximately 80-kDa, Endo H-resistant tyrosinase doublet, which represented mature glycoform of tyrosinase, was increased in melanocytes treated with Semen Cuscutae, and in those treated with Semen Persicae. Within those cells, tyrosinase was distributed outside ER resident protein calnexin. Conclusion Both Semen Cuscutae and Semen Persicae could induce tyrosinase maturation, stability and export from ER to distal site.