黄芩素对HaCaT细胞光老化模型ERK信号通路的影响

Effect of baicalein on ERK signaling pathways of HaCaT cell with light aging

目的研究黄芩素对皮肤光老化模型中细胞外调节蛋白激酶(ERK)信号通路的影响。方法使用照射强度为0.5 m J/cm2的UVB型紫外线辐照计,照射不同时间制备光老化细胞模型。细胞分为对照组,模型组,黄芩素1×10^(-7)、1×10^(-6)、1×10^(-5) mol/L组。使用流式细胞仪检测各组细胞中活性氧(ROS)量的变化,q RT-PCR检测各组细胞中白细胞介素-1α(IL-1α)、IL-8 m RNA表达量,蛋白免疫印迹法(Western blotting)检测各组细胞中IL-1α、IL-8、p-ERK、ERK蛋白的表达量。结果与对照组比较,模型组细胞出现皱缩、椭圆形,细胞边缘有破损现象,趋于死亡状态。与模型组相比,1×10^(-7) mol/L黄芩素组细胞破损现象减轻,1×10^(-6) mol/L黄芩素组细胞无明显的皱缩及破损现象,1×10^(-5) mol/L黄芩素组细胞状态良好,接近于正常状态;与对照组比较,模型组细胞中ROS的量显著升高(P<0.01)。与模型组比较,1×10^(-7)、1×10^(-6)、1×10^(-5) mol/L黄芩素组细胞中ROS的量显著降低(P<0.01);与对照组比较,模型组IL-1α、IL-8 m RNA表达量显著升高(P<0.01)。与模型组比较,1×10^(-7)、1×10^(-6)、1×10^(-5) mol/L黄芩素组细胞中IL-1α、IL-8 m RNA的表达量显著降低(P<0.01);与对照组比较,模型组细胞IL-1α、IL-8、p-ERK蛋白表达量显著升高(P<0.01),ERK蛋白表达量不变。与模型组比较,1×10^(-7)、1×10^(-6)、1×10^(-5) mol/L黄芩素组细胞中IL-1α、IL-8、p-ERK蛋白表达量显著降低(P<0.05、0.01),ERK蛋白表达量不变。结论黄芩素对HaCaT细胞光老化模型的保护作用主要通过降低ROS的量,阻断ERK信号通路,进而降低炎症因子的产生实现的。

Objective To study the effect of baicalein on the skin cell light aging and ERK signaling pathways. Methods Using UVB phototherapy device, of which light intensity was 0.5 mJ/cm^2, different exposure time was tried before establishing the best exposure time light aging model. The experiment can be divided into control group, model group and baicalein group（1 × 10^-7, 1 × 10^-6, 1 × 10^-5 mol/L） since the results of preliminary experiments shown that these three concentration of baicalein is the most effective concentration. Using flow cytometry instrument to test cells reactive oxygen species（ROS） content changes of each group. quantitative Real-time RCR（q RT-PCR） and Western blotting were respectively used to detect m RNA and protein expression in each group. Results The best exposure time for 5 min, and cell shrinkage, damage, apoptosis were observed in model group. Compared with model group, cell damage, shrinking phenomenon obviously reduced in 1 × 10^-7, 1 × 10^-6 mol/L baicalein group, and cell obviously tended to normal form in 1 × 10^-5 mol/L baicalein group. Compared with the control group, the ROS content of cells in model group was increased significantly（P〈0.01）. Compared with model group, the ROS content was decreased significantly in 1 × 10^-7, 1 × 10^-6, 1 × 10^-5 mol/L baicalein group（P〈0.01）; Compared with the control group, the expression of IL-1α and IL-8 m RNA was increased significantly（P〈0.01）, and the expression of IL-1-α, IL-8, and p-ERK protein was also increased significantly（P〈0.01） and ERK protein expression quantity remains the same in the model group. In 1 × 10^-7, 1 × 10^-6, 1 × 10^-5 mol/L baicalein group, compared with model group, IL-1α and IL-8 m RNA expression were decreased significantly（P〈0.01）, and IL-1-α, IL-8, and p-ERK protein expression were also decreased significantly（P〈0.05, 0.01） and ERK protein expression quantity remains the same. Conclusion Protection of baicalein on UVB induced cell light aging, mainly through reducing ROS content, blocking ERK signaling pathway and inhibiting inflammatory cytokines.