微波辐射诱导人神经母细胞瘤SH-SY5Y细胞凋亡的可能机制

J4 ›› 2011, Vol. 37 ›› Issue (3): 422-426.

微波辐射诱导人神经母细胞瘤SH-SY5Y细胞凋亡的可能机制

王程^1,2, 李琴³, 刘微¹, 闫岗林², 徐冶¹, 于洋¹, 刘师兵¹, 潘文干¹

（1.吉林医药学院生物化学教研室|吉林吉林132013； 2.吉林大学分子酶工程教育部重点实验室| 吉林长春130012；3 中国环境科学研究院环境标准研究所|北京100012）

收稿日期:2011-01-06 出版日期:2011-05-28 发布日期:2011-05-28
通讯作者: 王程 E-mail:E-mail:wangcheng10@mails.jlu.edu.cn
作者简介:王程（1979-）|男|辽宁省抚顺市人|讲师|在读医学博士|主要从事微波辐射损伤的防治研究。
基金资助:
吉林省科技厅白求恩基金资助课题（200705408）；吉林省教育厅科研基金资助课题（2009315）；吉林医药学院科研基金资助课题（200903）

Mechanism of apoptosis induced by microwave radiation in human neuroblastoma SH-SY5Y cells

WANG Cheng^1,2, LI Qin², LIU Wei¹, YAN Gang-Lin², XU Ye¹, XU Xiang¹, LIU Shi-Bing¹, BO Wen-An¹

(1. Department of Biochemistry,School of Basic Medical Sciences,Jilin Medical College,Jilin 132013,China;2. Key Laboratory for Molecular Enzymology and Engineering,Ministry of Education,Jilin University,Changchun 130023,China;3. Environmental Standards Institute,Chinese Research Academy of Environmental Sciences,Beijing 100012,China

Received:2011-01-06 Online:2011-05-28 Published:2011-05-28

摘要/Abstract

摘要：

目的：观察微波辐射对人神经母细胞瘤SH-SY5Y细胞凋亡的影响，并探讨其机制。方法：采用10、30和50 mW·cm^-2辐射强度的微波辐射SH-SY5Y细胞5 min，以假辐射组（0 mW·cm^-2）为对照，光镜下观察细胞形态变化；荧光显微镜下观察DAPI染色细胞核的形态；CTAB法提取细胞基因组DNA，电泳观察DNA ladder；台盼蓝拒染法检测细胞存活率；AnnexinV-FITC和PI双染色，流式细胞仪检测细胞凋亡率；MTT法检测细胞相对活性；蛋白质印迹检测细胞凋亡相关蛋白的表达。结果：微波辐射后SH-SY5Y细胞形态即刻改变，胞核、胞质结构欠清，细胞皱缩甚至脱壁；细胞核内的染色质出现不规则凝集，碎裂成2 ~ 5个微核；细胞DNA出现明显的梯状条带；细胞存活率随微波辐射强度增大逐渐降低，10 mW·cm^-2辐射组与假辐射组比较，细胞存活率差异无统计学意义（P>0.05），但30、50 mW·cm^-2辐射组细胞存活率明显低于假辐射组（P<0.05）；微波辐射后6 h的细胞凋亡率随辐射强度的增加逐渐升高，各辐射组细胞凋亡率均明显高于假辐射组（P<0.05）；微波辐射后24和48 h，细胞相对活力随辐射强度的增加明显降低，各辐射组的细胞相对活力均明显低于假辐射组（P<0.05）。微波辐射后，细胞凋亡相关蛋白Bcl-2、survivin表达水平逐渐下降，Bax、caspase-3　和caspase-7表达水平逐渐升高，caspase-8和-9表达水平无明显变化。结论：微波辐射可诱导SH-SY5Y细胞凋亡，其作用机制可能与survivin、Bcl-2蛋白表达水平下调、Bax蛋白表达水平上调及caspase-3和caspase-7的作用有关，而与caspase-8和caspase-9介导的caspase-3活化这一作用无密切关系。

关键词: 微波辐射；人神经母细胞瘤；SH-SY5Y细胞；细胞凋亡

Abstract:

Abstract:Objective
To observe the influence of microwave radiation in the apoptosis of human neuroblastoma SH-SY5Y cells and explore its mechanism. Methods Regarding 0 mW·cm^-2 as sham radiation group，the SH-SY5Y cells were radiated for 5 minutes by 10,30 and 50 mW·cm^-2 of microwave. The morphological changes of cells were observed under light microscope;the cell nucleus stained by DAPI were observed under fluorescence microscope;the cell genomic DNA was extracted by CTAB and electrophoresed to observe DNA ladder;the cell viability was detected by trypan blue exclusion assay;the apoptotic rate was detected by flow cytometry with annexin V-FITC/PI double staining;the relative activity of cells was detected by MTT assay;the expressions of apoptosis-related proteins were detected by Western blotting. Results After microwave radiation,the morphology of SH-SY5Y cells changed immediately,the nucleus and cytoplasm structure were less clear,and cell shrinkage or even off the wall were observed;chromatin in the nucleus appeared irregular condensation,and fragmentated into 2 to 5 micronuclei;cell DNA ladder was apparent. The survival rate of SH-SY5Y cells was reduced gradually with the increasing of microwave radiation intensity,there was no significant difference between 10 mW·cm^-2 radiation group and sham radiation group (P>0.05);however,the cell survival rates in 30 and 50 mW·cm^-2 radiation group were lower than that in sham radiation group(P<0.05). The apoptotic rate of SH-SY5Y cells was gradually increased with the microwave radiation intensity in 6 h after microwave radiation,and the apoptotic rates in each radiation group were higher than that in sham radiation group(P<0.05). Compared with sham radiation group,the cell relative activities of SH-SY5Y cells in each radiation group were markedly decreased 24 and 48 h after microwave radiation(P<0.05),and they were decreased with the increasing of microwave radiation intensity. After microwave radiation,the expressions of apoptosis related proteins Bcl-2 and survivin were decreased,the expressions of Bax,Caspase-3 and Caspase-7 were increased gradually,but the expressions of Caspase-8 and Caspase-9 changed little. Conclusion The apoptosis of SH-SY5Y cells can be induced by microwave radiation in a obvious dose-dependent manner,which may be related to the increase of Survivin and Bcl-2 proteins,the decrease of Bax protein and the action of the Caspase-3 and Caspase-7,but with little relationship of the activation of Caspase-3 mediated by caspase-8 and Caspase-9.

Key words: microwave radiation;human neuroblastoma, SH-SY5Y cells;apoptosis

中图分类号:

Q684

王程, 李琴, 刘微, 闫岗林, 徐冶, 于洋, 刘师兵, 潘文干. 微波辐射诱导人神经母细胞瘤SH-SY5Y细胞凋亡的可能机制[J]. J4, 2011, 37(3): 422-426.

WANG Cheng, LI Qin, LIU Wei, YAN Gang-Lin, XU Ye, XU Xiang, LIU Shi-Bing, BO Wen-An. Mechanism of apoptosis induced by microwave radiation in human neuroblastoma SH-SY5Y cells[J]. J4, 2011, 37(3): 422-426.