补骨脂异黄酮对UVB诱导HaCaT细胞凋亡的保护作用及机制研究

目的探讨补骨脂异黄酮对中波紫外线(UVB)诱导人永生化角质细胞(HaCaT)凋亡的保护作用及相关机制。方法采用四甲基噻唑蓝(MTT)法检测HaCaT细胞增殖率,筛选补骨脂异黄酮的实验浓度及UVB的照射时间,建立UVB诱导的HaCaT细胞凋亡模型。采用流式细胞术检测细胞凋亡率;试剂盒检测细胞中超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)、过氧化氢酶(CAT)活性;qRT-PCR法检测HaCaT细胞半胱氨酸蛋白酶(Caspase-3)mRNA表达;Western Blot法检测HaCaT细胞p-AKT、AKT、Caspase-3蛋白表达。结果选择10-5mol·L^-1浓度用于后续实验,10 min的UVB照射时间用于建立HaCaT细胞凋亡模型。与空白组比较,模型组的细胞增殖率明显下降,细胞凋亡率明显升高,SOD、GSH-Px及CAT活性显著降低,Caspase-3mRNA及p-AKT、Caspase-3蛋白表达显著上调,差异均有统计学意义(P<0.01)。与模型组比较,补骨脂异黄酮组的细胞增殖率显著升高,细胞凋亡率明显降低,SOD、GSH-Px、CAT活性显著升高,Caspase-3 mRNA及p-AKT、Caspase-3蛋白表达显著下调,差异均有统计学意义(P<0.05,P<0.01)。结论补骨脂异黄酮对UVB诱导的HaCaT细胞凋亡具有保护作用,可能与下调Caspase-3及p-AKT表达,提高抗氧化酶的活性有关。

Protective Effect and Mechanism of Psoralen Isoflavones on UVB-induced Apoptosis in HaCaT Cells

Objective To investigate the protective effect of psoralen isoflavones on the apoptosis of human permanent keratinocytes(HaCaT)cells induced by UVB and the related mechanism.Methods HaCaT cell proliferation rate was detected by tetramethylthiazole blue(MTT)method.The experimental concentration of psoralea isoflavones and UVB irradiation time were optimized,and the UVB-induced HaCaT cell apoptosis model was established.The apoptosis rate was detected by flow cytometry.The activities of superoxide dismutase(SOD),glutathione peroxidase(GSH-Px)and catalase(CAT)of the cells were detected by commercial kits.mRNA expression of cysteine protease(Caspase-3)in HaCaT cells was detected by qRT-PCR.Western Blot was used to detect the protein expression of p-AKT,AKT and Caspase-3 in HaCaT cells.Results A concentration of 10-5 mol·L^-1 was selected for application of psoralen isoflavones in subsequent experiments,and a duration of 10 min of UVB irradiation was used to establish the HaCaT cell apoptosis model.Compared with the control group,the cell proliferation rate and apoptosis rate of the model group were significantly decreased;activities of SOD,GSH-Px and CAT were significantly decreased;the relative expression level of Caspase-3 mRNA was significantly increased,and the relative expression levels of p-AKT and Caspase-3 were significantly increased,all with statistically significancet(P<0.01).Compared with the model group,the cell proliferation rate and apoptosis rate of the psoralen isoflavone group were significantly increased,SOD,GSH-Px and CAT activities were significantly increased;the relative expression level of Caspase-3 mRNA was significantly decreased,and the relative expression levels of p-AKT and Caspase-3 protein were significantly decreased,all with statistical significancet(P<0.05,P<0.01).Conclusion Psoralen isoflavones have a protective effect on UVB-induced HaCaT cell apoptosis,which may be related to the down-regulation of Caspase-3 and p-AKT expression and the enhancement of antioxidant enzyme activities.