加味当归芍药散抑制组蛋白乙酰化酶p300/血清环加氧酶-2通路调控核因子E2相关因子2在黄褐斑形成中的作用机制研究

The mechanism of modified Danggui Shaoyao powder inhibiting p300HAT/COX-2 pathway from regulating Nrf2 in the formation of chloasma

目的探讨加味当归芍药散抑制组蛋白乙酰化酶p300(p300HAT)/血清环加氧酶-2(COX-2)通路调控核因子E2相关因子2(Nrf2)在黄褐斑形成中的作用机制。方法健康人表皮黑素细胞及30只雌性小鼠纳入研究,采用不同浓度加味当归芍药散处理健康人表皮黑素细胞,肌肉注射黄体酮及紫外线照射建立黄褐斑小鼠模型,将小鼠随机分为空白组、低剂量组及高剂量组各10只,分别给予常规剂量0.9%氯化钠注射液、低剂量及高剂量加味当归芍药散处理,检测不同浓度处理下细胞活力、细胞黑素含量及细胞中p300HAT、COX-2及Nrf2的RNA及蛋白表达量,同时检测治疗前后3组小鼠皮肤及肝脏的p300HAT、COX-2及Nrf2的RNA、蛋白质表达量。结果加味当归芍药散浓度为1.0mg/mL时健康人表皮黑素细胞活力较对照组明显低,差异有统计学意义(P<0.05);加味当归芍药散处理的健康人表皮黑素细胞p300HAT、COX-2的RNA及蛋白表达量和黑素相对含量较对照组明显低,Nrf2的RNA及蛋白表达量较对照组明显高,且随着加味当归芍药散浓度增加,健康人表皮黑素细胞p300HAT、COX-2、Nrf2的RNA及蛋白表达量降低、升高越明显,健康人表皮黑素细胞黑素相对含量降低越明显,差异有统计学意义(P<0.05);治疗后仅低剂量、高剂量组p300HAT、COX-2的RNA、蛋白表达量较治疗前明显降低,Nrf2的RNA、蛋白表达量明显升高,且治疗后高剂量组各项指标变化较低剂量组明显,差异有统计学意义(P<0.05)。结论加味当归芍药散治疗黄褐斑的机制可能与其抑制p300HAT/COX-2通路调控Nrf2表达有关。

Objective To investigate the mechanism of modified Danggui Shaoyao powder inhibiting histone acetyltransferases p300 （p300HAT）/serum cyclooxygenase-2 （COX-2） pathway from regulating nuclear factor E2 related factor 2 （Nrf2） in the formation of chloasma .Methods The normal human melanocytes and 30 female mice were included in the study .The normal human epidermal melanocytes were treated with different concentra-tions of modified Danggui Shaoyao powder .The chloasma mouse models were established through intramuscular injection of progestin and ultraviolet irradiation .The mice were randomly divided into the blank control group ,low dose group and high dose group with 10 mice in each group .The three groups were treated with routine dose of normal saline ,low dose and high dose of modified Danggui Shaoyao powder ,respectively .The cell viability ,con-tent of cell melanin and expression levels of p300HAT ,COX-2 and Nrf2 RNA and protein in cells were detected after being treated with different concentrations .The expression levels of p300HAT ,COX-2 and Nrf2 RNA and protein in skin and liver of the three groups were also detected before and after treatment .Results When the con-centration of modified Danggui Shaoyao powder was 1 .0 mg/mL ,the activity of normal human melanocytes was significantly lower than that of the control group （ P 〈 0.05） .The expression levels of p300HAT and COX-2 RNA and protein in normal human melanocytes treated with modified Danggui Shaoyao powder were lower than those in control group .The expression levels of Nrf2 RNA and protein were significantly higher than those in con-trol group .With the increase of the concentration of modified Danggui Shaoyao powder ,the expression levels of p300HAT ,COX-2 and Nrf2 RNA and protein in normal human melanocytes decreased （ P 〈0.05） .After treat-ment ,the expression levels of 300HAT and COX-2 RNA and protein in the low dose group and high dose group were significantly lower than those before treatment ,while the expression levels of Nrf2 RNA and protein were significantly increased ,and the changes were more significant in the high dose group than the low dose group（ P〈0.05） .Conclusion The mechanism of modified Danggui Shaoyao powder in treating chloasma may be related to the inhibition of p300HAT/COX-2 pathway from regulating the expression of Nrf2 .