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目的:探讨灯盏细辛与赤芍配伍组方对H2O2诱导的PC12细胞氧化损伤的影响及其作用机制。方法:用H2O2(850μmol·L-1)处理PC12细胞6 h建立体外氧化应激模型,以不同质量浓度的灯盏细辛与赤芍配伍组方(6.25,12.5,25,50,100 mg·L-1)预处理PC12细胞(3×104~4×104个/mL)24 h,采用MTT法检测细胞活力,比色法测定细胞培养液中乳酸脱氢酶(LDH)活性、细胞裂解液中超氧化物歧化酶(SOD)活性和丙二醛(MDA)水平,利用荧光探针DCFH-DA和罗丹明123流式细胞术分别检测细胞内活性氧(ROS)和线粒体膜电位(Δψm)。结果:与正常对照组比较,模型组的细胞存活率下降至52.78%,细胞上清液中LDH释放量上升110.13 U·L-1,细胞内MDA和ROS水平显著增高,细胞内SOD和Δψm水平显下降(P<0.01);与模型组比较,灯盏细辛与赤芍配伍组方可明显增加细胞存活率,降低LDH活性,降低MDA水平,抑制细胞内活性氧的生成,增加SOD活性和使线粒体膜电位升高(P<0.05,P<0.01),且在一定范围呈剂量依赖性。结论:灯盏细辛与赤芍配伍组方对H2O2诱导的PC12细胞氧化损伤具有保护作用,其机制可能与其降低细胞脂质过氧化水平,提高抗氧化酶活力,抑制ROS生成和稳定线粒体膜电位有关。
OBJECTIVE: To investigate the effects of combination of Radix Astragali and Radix Astragali on the oxidative damage of PC12 cells induced by H2O2 and its mechanism. METHODS: The PC12 cells were treated with H2O2 (850 μmol·L-1) for 6 h to establish an in vitro oxidative stress model. The concentrations of different concentrations of Erigeron Breviscapus and Radix Astragali saponins were the same (6.25, 12.5, 25, 50, 100 mg·L-1). Pretreatment of PC12 cells (3×104~4×104 cells/mL) for 24 h. Cell viability was measured by MTT assay. Lactate dehydrogenase (LDH) activity in cell culture fluid and superoxide in cell lysate were measured by colorimetric method. Dismutase (SOD) activity and malondialdehyde (MDA) levels were measured using fluorescent probes DCFH-DA and rhodamine 123 flow cytometry to detect intracellular reactive oxygen species (ROS) and mitochondrial membrane potential (Δψm). RESULTS: Compared with the normal control group, the cell survival rate of the model group decreased to 52.78%, the LDH release in the cell supernatant increased by 110.13 U.L-1, the intracellular MDA and ROS levels increased significantly, and the intracellular SOD and Δψm levels increased. Significantly decreased (P<0.01); compared with the model group, the combination of Erigeron Breviscapus and Radix Paeoniae Alba can obviously increase cell survival rate, decrease LDH activity, decrease MDA level, inhibit the generation of reactive oxygen species, increase SOD activity and make The mitochondrial membrane potential increased (P<0.05, P<0.01) and was dose-dependent in a certain range. Conclusion: Combination of Erigeron Breviscapus and Radix Astragali has protective effects on H2O2-induced oxidative damage of PC12 cells, which may be related to the decrease of lipid peroxidation, increase of antioxidant enzyme activity, inhibition of ROS production and stabilization of mitochondrial membrane potential. .