白桦脂醇对A375细胞黑素合成及p38-MAPK信号通路调控机制的研究

Study on the Mechanism of Melanin Synthesis and p38-MAPK Signaling Pathway in A375 Cells by Betulin

目的:研究植物雌激素成分白桦脂醇对黑素合成中关键限速酶、MAPK信号通路调控作用。方法:使用NaOH裂解法、多巴醌氧化法检测黑素含量及酪氨酸酶活性,Western blot和PCR法测定MAPK通路中关键蛋白激酶和TYP、TRP-1、TRP-2、MITF、p38、p-p38蛋白含量及mRNA表达;加入p38信号通路阻断剂(SB203580)之后,测定上述蛋白和mRNA表达。结果:白桦脂醇对A375细胞黑素含量和酪氨酸酶活性有抑制作用(P<0.05或P<0.01),SB203580(10μmol/L)能阻断白桦脂醇对A375细胞黑素含量和酪氨酸酶活性抑制作用;白桦脂醇对A375细胞中上述蛋白及mRNA表达有抑制作用(P<0.05或P<0.01),加入SB203580后,阻断p38-MAPK信号通路,白桦脂醇以上作用受到显著抑制(P<0.05或P<0.01)。结论:表明白桦脂醇通过MAPK信号通路中的p38-MAPK信号通路抑制A375细胞黑素合成。

Objective:To study the regulation of key rate-limiting enzymes and MAPK signaling pathway in the synthesis of melanin by the phytoestrogenic component-betulin.Methods:The melanin content and tyrosinase activity were measured by NaOH lysis method and dopaquinone oxidation method,and the key protein kinases in the MAPK pathway and the protein contents and mRNA expressions of TYP,TRP-1,TRP-2,MITF,p38 and p-p38 were determined by Western blot and PCR method.After adding the p38 signaling pathway blocker(SB203580),the above proteins and related mRNA expressions were determined.Results:Betulin inhibited melanin content and tyrosinase activity in A375 cells(P<0.05 or P<0.01),SB203580(10 μmol/L)could block the inhibitory effect of betulin on melanin and tyrosinase activity of A375 cells;Betulin could inhibite the above proteins and mRNA expressions in A375 cells(P<0.05 or P<0.01).After adding SB203580,it blocked the p38-MAPK signaling pathway and above effects of betulin were inhibited(P<0.05 or P<0.01).Conclusion:Betulin inhibits melanin synthesis in A375 cells via the p38-MAPK signaling pathway which is in the MAPK signaling pathway.