The impact of α-particles irradiation on src kinase activity and autophagy system mediated by ROS / 中华放射医学与防护杂志

ABSTRACT

Objective To investigate the modulation role of autophagy in radiation-induced cell death by detecting the response of src kinase activity and autophagy in HEK293 cells irradiated with different dose of α-particle.Methods HEK293 cells were irradiated by contral group (0 cGy) a low dose group (10 cGy) and high dose group (300 cGy) α-particles.Molecular probe 2',7'-dichlorofluorescin (DCFHDA) was used to detect the cell ROS.The src kinase activity and endogenous protein level of LC3Ⅰ/Ⅱ were monitored by Western Blot.Cell death rate of irradiated cells pretreated with autophagy inducer of rapamycin was tested by flow cytometry.Results Compared with control group,the ratio of LC3Ⅰ/Ⅱ decreased (t =4.07,P ＜ 0.05) and the percentage of cells with GFP-LC3 punctuate dots increased (t =12.29,P ＜0.05) under 10 cGy irradiation,indicating the induction of autophagy.On the contrary,the ratio of LC3Ⅰ/Ⅱ increased (t =2.93,P ＜ 0.05) and the GFP-LC3 morphology had no obvious change under 300 cGy irradiation.The cellular ROS level reached to the maximum value at 4 h postirradiation.Both 10 cGy and 300 cGy irradiation could elevate the ROS level (t =17.93,22.88,P ＜0.05),whereas the amplitude of elevation of 300 cGy irradiation was higher than that of 10 cGy irradiation (t =15.76,22.66,14.22,P ＜ 0.05).Compared with control group,the 419th site of tyrosine residue in src kinase manifested hyper-phosphorylation (t =5.66,P ＜0.05) under 10 cGy irradiation whereas it had hypo-phosphorylation under 300 cGy irradiation (t =4.67,P ＜ 0.05).Treatment of cells with DMSO could partly restore the impact on src kinase activity under high or low dose irradiation.Pre-treating the cells with autophagy inducer rapamycin could reduce cell death under 300 cGy irradiations (t =12.14,P ＜ 0.05).Conclusions High or low dose of α-particles irradiation could inhibit or activate src kinase and autophagy system,respectively.ROS mediated the response of src kinase activity and autophagy system induced by irradiation.Modulation of autophagy could desensitize cell responses to irradiation.