脱氧熊果苷和氢醌对人黑素小体结构完整性及Pmel17蛋白表达影响的差异研究

Comparison of effects of deoxy-arbutin and hydroquinone on structural integrity of melanosomes and expression of Pmel17 in human melanocyte

目的探讨脱氧熊果苷和氢醌对人黑素小体结构完整性和前黑素小体蛋白17(Pmel17)表达影响的差异。方法体外分离并纯化黑素瘤细胞系MNT1中黑素小体,实验分3组,对照组、氢醌组、脱氧熊果苷组分别加入磷酸盐缓冲液(PBS)、10μmol/L氢醌、100μmol/L脱氧熊果苷,37℃孵育30 min,通过透射电镜观察各组黑素小体超微结构的改变。体外培养MNT1细胞,实验分3组,对照组、氢醌组、脱氧熊果苷组分别加入PBS、10μmol/L氢醌、100μmol/L脱氧熊果苷,37℃孵育24 h,蛋白免疫印迹法检测各组Pmel17蛋白的表达水平。采用单因素方差分析和Bonferroni法比较组间差异。结果对照组黑素小体超微结构完整,黑素小体破坏率为(14.80±3.70)%,Pmel17蛋白相对表达量为0.84±0.04;氢醌组黑素小体膜结构破坏,黑素小体裂解,黑素小体破坏率为(54.40±3.65)%,Pmel17相对表达量为0.61±0.03;脱氧熊果苷组黑素小体膜结构也遭破坏,部分黑素小体裂解,黑素小体破坏率仅为(27.60±3.65)%,Pmel17蛋白相对表达量为0.49±0.03。氢醌组与对照组、脱氧熊果苷组与对照组、脱氧熊果苷组与氢醌组在黑素小体结构完整性和Pmel17蛋白表达量比较差异均有统计学意义(P均<0.05)。结论在体外实验中,与氢醌相比,脱氧熊果苷仅轻度破坏黑素小体超微结构,但更明显抑制了Pmel17蛋白的表达。

Objective To compare the effects between deoxy-arbutin and hydroquinone on the structural integrity and the expression of Pmel 17 protein of human melanosome. Methods Melanosomes were isolated and purified in vitro from melanoma cell line MNT1 and divided into the control, hydroquinone and deoxy-arbutin groups. In each group, PBS, 10 μmol/L hydroquinone and 100 μmol/L deoxy-arbutin were supplemented and incubated at 37℃ for 30 min. The ultrastructure of melanosomes in each group was observed by transmission electron microscopy. MNT1 cells were cultured in vitro and divided into three groups: control group, hydroquinone group and deoxy-arbutin group supplemented with PBS, 10 μmol/L hydroquinone and 100 μmol/L deoxy-arbutin, respectively. The cells were incubated at 37 ℃ for 24 h. The expression level of Pmel17 protein in each group was detected by Western blot. One-way ANOVA analysis and Bonferroni’s multiple comparisons test were used for comparison among different groups. Results In the control group, the ultrastructure of melanosomes was intact, the destruction rate of melanosomes was(14.80±3.70)%, and the relative expression level of Pmel17 protein was 0.84±0.04. In the hydroquinone treatment group, the membrane structure of melanosomes was destroyed and melanosomes were disrupted. The destruction rate of melanosomes was(54.40±3.65)%, and the relative expression level of Pmel17 protein was 0.61±0.03.In the deoxy-arbutin treatment group, the membrane structure of melanosomes was also destroyed, and some melanosomes were disrupted. The destruction rate of melanosomes was only(27.60±3.65)%, and the relative expression level of Pmel17 protein was 0.49±0.03. There was significant difference in melanosome structural integrity and Pmel17 protein expression between any two groups(all P < 0.05). Conclusions Compared with hydroquinone, deoxy-arbutin can cause less damage to the ultrastructure of melanosomes, but more significantly down-regulate the expression level of Pmel17 protein in vitro.