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目的:探讨延胡索乙素(THP)对放射所致血管内皮细胞损伤的保护作用及其机制。方法:以THP预处理人血管内皮细胞株EA.hy926后,以10 Gyγ射线照射,建立放射性细胞损伤模型。光镜及电镜观察细胞形态改变,流式细胞仪检测细胞凋亡率,分光光度计检测细胞照射前后乳酸脱氢酶(LDH)、丙二醛(MDA)的含量变化,流式细胞仪检测细胞内活性氧(ROS)生成,蛋白质印迹法检测细胞色素C蛋白表达,酶标仪检测Caspase-3的活性变化。结果:细胞经照射后,细胞形态及超微结构改变均显示典型的凋亡改变,而THP预处理组的细胞凋亡改变明显轻于其他实验组。流式检测结果示,单纯照射组血管内皮细胞凋亡率为(75.35±7.55)%,而THP+R处理组明显降低,最低达(32.05±4.15)%,P<0.01;单纯照射组血管内皮细胞脂质氧化损伤指标LDH、MDA明显升高,而低中高剂量的THP+R处理组明显降低,P<0.01。单纯照射组血管内皮细胞内的ROS含量及Caspase-3的活性亦明显升高,而THP+R组细胞内的ROS含量及Caspase-3的活性明显降低,P<0.01。同时,THP+R组细胞内细胞色素C蛋白表达均显著低于单纯照射组,P<0.05。结论:THP可能通过抑制放射诱导ROS生成,抑制细胞色素C蛋白、Caspase-3激酶激活,抑制细胞内脂质氧化损伤等作用,防治放射所致的内皮细胞损伤,这可能为预防或减轻放射治疗所致的肺损伤提供一些新思路。
Objective: To investigate the protective effect of tetrahydropalmatine (THP) on the injury of vascular endothelial cells induced by radiation and its mechanism. Methods: The THP-pretreated human vascular endothelial cell line EA.hy926 was irradiated with 10 Gy γ-rays to establish a model of radiation-induced cell injury. The morphological changes of cells were observed under light microscope and electron microscope. The apoptosis rate was detected by flow cytometry. The contents of lactate dehydrogenase (LDH) and malondialdehyde (MDA) in cells before and after irradiation were measured by spectrophotometer. Flow cytometry The generation of reactive oxygen species (ROS), the expression of cytochrome C protein by Western blotting, and the activity change of Caspase-3 by microplate reader. Results: After the cells were irradiated, the changes of cell morphology and ultrastructure showed the typical changes of apoptosis. However, the apoptosis of THP pretreatment group was obviously lighter than that of other experimental groups. The results of flow cytometry showed that the apoptotic rate of vascular endothelial cells in pure irradiation group was (75.35 ± 7.55)%, while that in THP + R group was the lowest (32.05 ± 4.15)%, P <0.01; Lipid oxidative damage indicators LDH, MDA were significantly increased, while low, medium and high doses of THP + R treatment group was significantly reduced, P <0.01. The content of ROS and the activity of Caspase-3 in vascular endothelial cells of pure irradiation group were significantly increased, while the content of ROS and the activity of Caspase-3 in THP + R group were significantly decreased (P <0.01). Meanwhile, the intracellular cytochrome C protein expression in THP + R group was significantly lower than that in the pure irradiation group, P <0.05. CONCLUSION: THP may prevent and reduce the damage of endothelial cells induced by radiation by inhibiting the radiosensitization of ROS, inhibiting the activation of cytochrome C protein and Caspase-3 kinase and inhibiting the lipid oxidation injury in the cells, which may be the prevention or reduction of radiotherapy The resulting lung injury provides some new ideas.