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目的探讨乙酰化白藜芦醇对长波紫外线(UVA)照射后人永生化角质形成细胞HaCaT内核转录相关因子NRF2(NF-E2-related factors 2)活化和活性氧(reactive oxygen species,ROS)水平的影响。方法用CCK-8试剂盒检测不同剂量(0、5、10、20、30、40、60、80 J/cm~2)UVA照射后24 h,及不同浓度(0、0.5、1、2μmol/L)乙酰化白藜芦醇干预24 h后再经20 J/cm~2 UVA照射后24 h HaCaT细胞的存活率;Western blot检测20 J/cm~2 UVA照射HaCaT细胞后不同时间点(0、3、6、12、24、48 h)及2μmol/L乙酰化白藜芦醇干预24 h后再经20 J/cm~2 UVA照射后6 h细胞核内NRF2蛋白表达的变化;Annexin V-FITC/PI流式细胞术检测2μmol/L乙酰化白藜芦醇干预HaCaT细胞24 h后再经20 J/cm~2 UVA照射后24 h细胞的凋亡率和死亡率;二氯荧光黄二乙酸酯(dichlorodihydrofluorescein diacetate,DCFH-DA)荧光探针标记细胞检测HaCaT细胞中ROS水平的变化。结果 10~80 J/cm~2 UVA照射后24 h,HaCaT细胞存活率随照射剂量的增加逐渐降低(P<0.05);20 J/cm~2 UVA照射HaCaT细胞后0 h,细胞核内NRF2的表达急剧增强(P<0.01),照射后3~48 h又逐渐减弱(P<0.05)。乙酰化白藜芦醇预处理能显著提高UVA照射后细胞的存活率(P<0.05),降低其凋亡率和死亡率(P<0.05),促进细胞核内NRF2蛋白的表达(P<0.01),并降低细胞内ROS水平(P<0.01)。结论乙酰化白藜芦醇能减轻UVA照射引起的HaCaT细胞增殖抑制、凋亡和死亡,其机制可能与其促进NRF2蛋白活化,降低细胞内ROS水平,从而减轻UVA照射对细胞的氧化损伤有关。
Objective To investigate the activation of NF-E2-related factors 2 and the level of reactive oxygen species (ROS) in human immortalized keratinocytes after acetylated resveratrol exposure to UVA. influences. Methods CCK-8 kit was used to detect different doses (0, 5, 10, 20, 30, 40, 60, 80 J/cm~2) of UVA irradiation for 24 h, and different concentrations (0, 0.5, 1, 2 μmol/ml). L) The survival rate of HaCaT cells 24 h after 20 J/cm~2 UVA irradiation after acetylated resveratrol treatment for 24 h; Western blot detection of 20 J/cm~2 UVA irradiated HaCaT cells at different time points (0 (3, 6, 12, 24, 48 h) and 2 μmol/L acetylated resveratrol interfered with NRF2 protein expression in the nucleus 6 h after irradiation with 20 J/cm~2 UVA for 24 h; Annexin V- FITC/PI flow cytometry was used to detect the apoptosis rate and mortality of cells treated with 2 μmol/L acetylated resveratrol for 24 h after irradiation with 20 J/cm~2 UVA for 24 h. Dichlorodihydrofluorescein diacetate (DCFH-DA) fluorescent probe cells were used to detect the changes of ROS level in HaCaT cells. Results After 10 to 80 J/cm~2 UVA irradiation for 24 h, the survival rate of HaCaT cells gradually decreased with the increase of irradiation dose (P<0.05); after 20 J/cm~2 UVA irradiation of HaCaT cells, 0 h, the nuclei of NRF2 The expression increased sharply (P<0.01) and gradually decreased after 3 to 48 h (P<0.05). Acetylated resveratrol pretreatment could significantly increase the survival rate of cells irradiated with UVA (P<0.05), decrease the rate of apoptosis and mortality (P<0.05), and promote the expression of NRF2 protein in the nucleus (P<0.01). , and reduce intracellular ROS levels (P < 0.01). Conclusion The acetylated resveratrol can reduce the proliferation inhibition, apoptosis and death of HaCaT cells induced by UVA irradiation. Its mechanism may be related to promoting the activation of NRF2 protein and reducing the intracellular ROS level, thereby reducing the oxidative damage of cells induced by UVA irradiation.