小分子化合物I942对黑素细胞黑素合成的促进作用

Promoting effect of small molecule compound I942 on melanogenesis in melanocytes

目的研究小分子化合物I942对黑素细胞黑素合成的作用及相关机制。方法采用多巴染色法对正常人永生化黑素细胞系PIG1进行鉴定,用不同浓度的小分子化合物I942处理黑素细胞PIG1。用CCK-8法检测小分子化合物I942对细胞活力的影响,用氢氧化钠裂解法检测细胞中黑素含量,用多巴氧化反应法检测细胞中酪氨酸酶活性,用qRT-PCR检测细胞中黑素合成相关蛋白小眼畸形相关转录因子(MITF)、酪氨酸酶(TYR)、酪氨酸酶相关蛋白1(TRP1)、酪氨酸酶相关蛋白2(TRP2)mRNA的表达量,用蛋白质印迹法检测TYR、TRP2的蛋白表达情况。结果在0~50μmol/L的浓度范围内,小分子化合物I942对黑素细胞PIG1细胞活力的影响差异无统计学意义(P>0.05)。小分子化合物I942可增加黑素细胞PIG1中黑素含量(P<0.01),升高细胞内酪氨酸酶活性(P<0.01)。使用小分子化合物I942处理后,黑素细胞PIG1中MITF、TYR、TRP1、TRP2 mRNA表达均上升(P<0.01,P<0.05),TYR、TRP2蛋白表达无明显变化。结论小分子化合物I942在对细胞活力无明显抑制作用的基础上,可通过升高酪氨酸酶活性增加黑素细胞PIG1中黑素含量,同时黑素合成相关蛋白MITF、TYR、TRP1、TRP2的mRNA表达也升高。

Objective To explore the effect of small molecule compound I942 on melanogenesis in melanocytes and the possible mechanism.Methods Dopa staining was used to identify the PIG1 normal human immortal melanocyte cell line.PIG1 melanocytes were treated with different concentrations of small molecule compound I942.The effect of small molecule compound I942 on the cell viability of PIG1 melanocytes was detected by CCK-8 assay.The content of melanin was determined by sodium hydroxide solubilization.The activity of tyrosinase was detected by dopa oxidation.The mRNA expression of melanin synthesis-related proteins(smicrophthalmia-associated transcription factor[MITF],tyrosinase[TYR],tyrosinase-related protein 1[TRP1]and tyrosinase-related protein 2[TRP2])were detected by qRT-PCR.The protein expression of TYR and TRP2 was detected by Western blotting.Results There were no significant differences in the effect of small molecule compound I942 on the cell viability of PIG1 melanocytes at the concentration ranging from 0μmol/L to 50μmol/L.Small molecule compound I942 was found to increase both melanin content and tyrosinase activity in PIG1 melanocytes(both P<0.01).The mRNA levels of MITF,TYR,TRP1 and TRP2 were also increased after being treated with small molecule compound I942(P<0.01,P<0.05).There were no significant changes in the expression of TYR or TRP2 protein.Conclusion Small molecule compound I942 can increase the melanin content of PIG1 melanocytes by increasing the tyrosinase activity with no obvious inhibition of cell viability.The mRNA expression of melanin synthesis-related proteins(MITF,TYR,TRP1 and TRP2)are also increased.