摘要
目的:探讨鸦胆子苦醇(Brusatol,BRU)对人非小细胞肺癌H1299细胞辐射增敏作用及其机制。方法:利用CCK-8法检测BRU对H1299细胞的抑制率和半数抑制浓度(IC50),将H1299细胞分为对照组、BRU组(IC50)、X射线组(2 Gy)和BRU联合X射线组(BRU+2 Gy)。应用克隆形成实验检测细胞存活率,流式细胞仪检测照射后24 h细胞凋亡率和胞内活性氧含量(reactive oxygen species,ROS);Western blotting技术检测PI3K/Akt/Nrf2信号通路和线粒体凋亡通路相关蛋白表达;免疫荧光技术实现Nrf2蛋白定位。结果:BRU的IC50为100 nmol/L,用该浓度处理H1299细胞24 h后,Nrf2表达量最低;100 nmol/L BRU联合2 Gy的X射线照射后,与对照组比较,BRU组、2 Gy组和BRU+2 Gy组细胞存活率降低、细胞凋亡率和ROS升高;与2 Gy组比较,BRU+2 Gy组细胞存活率降低、细胞凋亡率和ROS升高;与对照组比较,BRU组、2 Gy组和BRU+2 Gy组PI3K和Akt磷酸化水平降低、Nrf2蛋白表达量降低,细胞核中Nrf2蛋白含量减少,线粒体凋亡通路相关蛋白表达量升高;与2 Gy组比较,BRU+2 Gy组PI3K和Akt磷酸化水平降低、线粒体凋亡通路相关蛋白表达量升高。结论:BRU联合X射线通过抑制PI3K/Akt/Nrf2信号通路,增加H1299细胞凋亡,从而增强H1299细胞的辐射敏感性。
Objective:To discuss the radiosensitizing effects of brusatol(BRU)on human non small cell lung cancer H1299 cells and its mechanism.Methods:CCK-8 method was used to detect the inhibitory rate and IC50 of BRU on H1299 cells,H1299 cells were divided into the control group,BRU group(IC50),X-ray group(2 Gy),BRU and X-ray group(BRU+2 Gy).Colony formation assay was used to detect the cell survival rate,and flow cytometry was used to detect the apoptosis rate within 24 h and the contents of intracellular reactive oxygen species(ROS)after irradiation;Western blotting technique was used to detect the expressions of PI3K/Akt/Nrf2 signaling pathway and mitochondrial apoptosis pathway related proteins;immunofluorescence to realize the localization of Nrf2 protein.Results:IC50 of BRU was 100 nmol/L,the expressions of Nrf2 were the lowest after dealing H1299 cells with the concentration for 24 hours;After dealing H1299 cells with 100nmol/L combined with 2 Gy X-ray irradiation,compared with the control group,the cell survival rate lowered,cellular apoptosis rate and ROS rose in BRU group,2 Gy group and BRU+2 Gy group;compared with 2 Gy group,the cell survival rate reduced,cellular apoptosis rate and ROS rose in BRU+2 Gy group;compared with the control group,phosphorylation levels of PI3K and Akt decreased,the expressions of Nrf2 protein lowered,the contents of Nrf2 protein in the nucleus reduced,the expressions of mitochondrial apoptosis pathway related proteins increased in BRU group,2 Gy group and BRU+2 Gy group;compared with 2 Gy group,phosphorylation levels of PI3K and Akt decreased,the expressions of mitochondrial apoptosis pathway related proteins increased in BRU+2 Gy group.Conclusion:BRU and X-ray could enhance the radiosensitivity of H1299 cells via inhibiting PI3K/Akt/Nrf2 signaling pathway and increasing the apoptosis of H1299 cells.
作者
张芳
杨蓉佳
付朝霞
ZHANG Fang;YANG Rongjia;FU Zhaoxia(Huachi County People’s Hospital,Huachi 745600,China;Gansu Provincial Hospital)
出处
《西部中医药》
2021年第7期38-43,共6页
Western Journal of Traditional Chinese Medicine
基金
甘肃省人民医院特色建设自选课题(201806)。
