Wang Yu-Rong, Yang Kang, Cui Wei-Ye, Piao Xiang-Lan
School of Pharmacy, Minzu University of China, Beijing 100081, China.
Zhongguo Zhong Yao Za Zhi. 2018 Mar;43(5):1014-1020. doi: 10.19540/j.cnki.cjcmm.2018.0034.
This study focuses on the therapeutical effect of flavonoids from Gynostemma pentaphyllum on human lung carcinoma A549 cells induced by H₂O₂ oxidative stress and its possible mechanisms. The oxidative damage model was established using different concentrations H₂O₂ to induce A549 cell for different hours, and then treated with the flavonoids for 10 hours. The effects of flavonoids from G. pentaphyllum on cell viability of A549 cell damaged by H₂O₂ were detected by MTT assay. The contents of ROS were detected by DCFH-DA fluorescent probe method via flow cytometer. The contents of MDA, SOD and GSH were detected by TBA，NBT and DTNB-linked colorimetry assay, respectively. Expressions levels of Nrf2, NQO1 and HO-1 in A549 cells were evaluated by Western blot. The results showed that the cell activity was decreasing with the rise of H₂O₂ concentration within the range of 200-700 μmol·L⁻¹. The cell viability was 60.4% after treated with 500 μmol·L⁻¹H₂O₂ for 10 h, so it was chosen to be as an oxidant stress model. Compared with normal group，the contents of SOD, GSH and HO-1 expressions were lower after damaged with H₂O₂. On the contrary, the contents of ROS and MDA expressions were increased. Compared with model group, the contents of SOD, GSH and the expressions of Nrf2, NQO1 and HO-1 were increased after treated with flavonoids from G. pentaphyllum. The above results demonstrate that flavonoids from G. pentaphyllum may attenuate the effect of H₂O₂-induced oxidative stress on A549 cell by resisting oxidation. The finding may provide a biological evidence for the application of the G. pentaphyllum to fight the oxidative stress related diseases.
本研究聚焦于绞股蓝总黄酮对H₂O₂氧化应激诱导的人肺癌A549细胞的治疗作用及其可能机制。采用不同浓度H₂O₂诱导A549细胞不同时长建立氧化损伤模型，随后用总黄酮处理10小时。通过MTT法检测绞股蓝总黄酮对H₂O₂损伤的A549细胞活力的影响。采用DCFH - DA荧光探针法经流式细胞仪检测ROS含量。分别采用TBA、NBT和DTNB比色法检测MDA、SOD和GSH含量。通过蛋白质免疫印迹法评估A549细胞中Nrf2、NQO1和HO - 1的表达水平。结果显示，在200 - 700 μmol·L⁻¹范围内，随着H₂O₂浓度升高细胞活性降低。用500 μmol·L⁻¹H₂O₂处理10小时后细胞活力为60.4%，故选择此作为氧化应激模型。与正常组相比，H₂O₂损伤后SOD、GSH含量及HO - 1表达降低。相反，ROS含量及MDA表达升高。与模型组相比，绞股蓝总黄酮处理后SOD、GSH含量及Nrf2、NQO1和HO - 1表达升高。上述结果表明，绞股蓝总黄酮可能通过抗氧化减轻H₂O₂诱导的氧化应激对A549细胞的影响。该发现可能为绞股蓝用于对抗氧化应激相关疾病提供生物学依据。
