紫铆花素对黑色素瘤细胞与角质形成细胞共培养体系中黑色素合成的影响

Effects of Butein on Melanosome Synthesis in the Co-culture System of Keratinocytes and Melanoma Cells

目的探讨不同浓度紫铆花素对黑素瘤细胞A375与角质形成细胞Hacat共培养体系细胞增殖、酪氨酸酶(TYR)活性、黑色素合成及酪氨酸相关蛋白1(TRP-1)、酪氨酸相关蛋白2(TRP-2) mRNA表达的影响。方法建立黑色素瘤细胞A375与角质形成细胞Hacat共培养体系,以不同浓度紫铆花素处理共培养体系48 h,采用噻唑蓝(MTT)法测定细胞增殖,氢氧化钠(Na OH)裂解法测定细胞黑色素含量,多巴氧化速率法测定黑色素细胞TYR活性,实时荧光聚合酶链反应(Real-time PCR)法检测TRP-1、TRP-2基因表达。结果与细胞共培养体系(即阴性对照组)比较,1.0,5.0μg·mL^(-1)紫铆花素与共培养体系细胞共孵育48 h后,可明显促进共培养细胞增殖(P<0.05或P<0.01),促进黑色素细胞TYR活性(P<0.05或P<0.01);在0.5～50μg·mL^(-1)范围内,紫铆花素可促进共培养体系黑色素合成(P<0.05或P<0.01);在0.1～0.5μg·mL^(-1)范围内,紫铆花素各剂量组TRP-1 mRNA表达升高(P<0.05,P<0.01);在0.5～5.0μg·mL^(-1)范围内,紫铆花素各剂量组TRP-2 mRNA表达升高(P<0.01)。讨论紫铆花素可能通过促进TYR活性和TRP-1、TRP-2mRNA表达增强共培养体系黑色素合成功能。

Objective To investigate the effects of butein on the proliferation, tyrosinase (TYR) activity, melanin synthesis and the mRNA expression of tyrosine-related protein 1 (TRP-1) and tyrosine-related protein 2(TRP-2) in the in vitro co-culture model of keratinocytes and melanoma cells. Methods The in vitro co-culture model of A375 and Hacat cells was established.Melanosome synthesis changes induced by different concentration of butein in mixed cultures of A375 and Hacat cells were observed 48 h after treatment, respectively.The cell proliferation rate was detected by MTT, the melanin content was measured with a sodium hydroxide solubilization method, the activity of TYR was estimated by measuring the rate of oxidation of L -dopa, and the mRNA expression of TRP-1 and TRP-2 were determined by real-time PCR. Results Compared with negative control group, after the treatment with butein (1.0 μg·mL -1 and 5.0 μg·mL -1 ) for 48 h, the cell proliferation rate of co-culture system of A375 and Hacat cells were significantly increased ( P <0.05 or P <0.01), the content of melanin and TYR activity was increased ( P <0.05 or P <0.01), the mRNA expression of TRP-1 and TRP-2 was increased ( P <0.01). Conclusion The butein up-regulates TYR activity, as well as the mRNA expression of TRP-1 and TRP-2, and then promotes melanosome synthesis in co-culture system of A375 and Hacat cells.