| 范华东,孙宇翔,陈少鹏,许绍海,吴李君.α粒子辐射对ROS介导src激酶活性和自噬的影响[J].中华放射医学与防护杂志,2015,35(7):481-484,527 |
| α粒子辐射对ROS介导src激酶活性和自噬的影响 |
| The impact of α-particles irradiation on src kinase activity and autophagy system mediated by ROS |
| 投稿时间：2015-02-16 |
| DOI：10.3760/cma.j.issn.0254-5098.2015.07.001 |
| 中文关键词: α粒子 细胞自噬 活性氧 src激酶 |
| 英文关键词:α-particles Autophagy Reactive oxygen species src kinase |
| 基金项目:国家自然科学基金(81273004,31470829) |
|
| 摘要点击次数: 2864 |
| 全文下载次数: 2246 |
| 中文摘要: |
| 目的 通过检测人胚肾上皮细胞HEK293受不同剂量α粒子照射后,src激酶活性和细胞自噬系统的变化,探讨调控细胞自噬对高剂量辐射诱导细胞死亡的影响。方法 将人胚肾上皮细胞HEK293分为3组:0 cGy(对照)组、241Am α粒子照射低剂量(10 cGy)组和高剂量(300 cGy)组。应用免疫印迹实验检测细胞内源性蛋白LC3I/II和src激酶的变化;分子探针检测细胞内活性氧(ROS)水平;用ROS淬灭剂DMSO预处理照射细胞,并用免疫印迹法检测照射后src激酶的变化;使用自噬诱导剂雷帕霉素预处理照射细胞,以PI染色流式细胞术测定细胞死亡率。结果 与0 cGy相比,10 cGy α射线照射后LC3I/II比例降低(t=4.07,P<0.05),胞质内具有绿色荧光点状GFP-LC3的细胞比例升高(t=12.29,P<0.05),自噬被诱导;而300 cGy照射后,LC3I/II比例升高(t=2.93,P<0.05),胞质内GFP-LC3形态无明显变化,自噬被抑制。与0 cGy相比,10和300 cGy照射后4 h均能提升细胞内ROS水平(t=17.93、22.88,P<0.05),且300 cGy比10 cGy照射诱发的ROS更多(t=15.76、22.66、14.22,P<0.05)。与0 cGy相比,10 cGy照射使src激酶419位酪氨酸磷酸化水平升高(t=5.66,P<0.05),而300 cGy照射则降低其磷酸化水平(t=4.67,P<0.05)。DMSO能够部分逆转高低剂量照射对src激酶活性的影响。辐照前以自噬诱导剂雷帕霉素处理细胞则能降低300 cGy照射诱发的细胞死亡率(t=12.14,P<0.05)。结论 高低剂量α粒子分别抑制和激活src激酶以及细胞自噬,ROS参与介导辐照对src激酶活性及自噬系统的影响。对自噬系统的干预能够降低细胞的辐射敏感性。 |
| 英文摘要: |
| 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 post-irradiation. 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. |
| HTML 查看全文 查看/发表评论 下载PDF阅读器 |
| 关闭 |
|
|
|