【作者】 杨明建；

【导师】 展永； 赵葵；

【作者基本信息】 河北工业大学 ， 生物物理， 2007， 硕士

【摘要】 随着质子和重离子治疗肿瘤的应用日益为人们所接受,高传能线密度(LET)辐射哺乳动物细胞的生物学效应机理研究越来越受到重视。本研究利用琼脂糖凝胶电泳配合先进的成像系统技术、集落形成实验等传统的细胞生物学分析手段以及基因芯片这一现代分子生物学实验技术研究了辐射致质粒DNA和一些细胞的生物学效应,以期为重离子和质子治癌、硼中子俘获治疗(BNCT)、太空辐射和核辐射的危险性评估及其防护等提供有价值的实验依据。实验过程及初步结果如下:利用60Co-γ射线对不含和含自由基清除剂的水溶液pUC19质粒DNA进行辐照,使用凝胶电泳方法分析实验样品,结果表明在水状DNA溶液环境下,γ射线辐射过程中产生的自由基是诱发DNA单链和双链断裂的重要原因;甘露醇,维生素C和茶多酚是很好的自由基清除剂,在相同条件下甘露醇比维生素C具有更强的防护作用;对比以前做过的重离子辐照实验,发现自由基清除剂对γ射线产生自由基的清除作用更加明显,这对于评估重离子辐照中自由基的作用具有十分重要的参考价值。利用60Co-γ射线和238Puα源发射的α粒子对U251神经胶质瘤细胞和HepG2肝癌细胞进行辐照,结果表明:U251细胞比HepG2细胞具有更高的辐射敏感性;α粒子比γ射线对细胞增殖的抑制作用更加明显;在同等剂量下,α粒子与γ射线相比具有更高的相对生物学效应(RBE)。在兰州近代物理研究所重离子研究装置(HIRFL)辐照终端,用5Gy能量为80MeV/u的12C6+离子对L02正常肝细胞进行了辐照,用22K的人类全基因组寡核苷酸芯片分析基因转录谱,结果表明辐照后培养6小时细胞发生差异表达的基因有37个,其中19个基因表达下调,18个基因表达上调。辐照后培养24小时细胞发生差异表达的基因有269个,202个基因表达下调,67个基因表达上调。这些基因大多与细胞信号转导、细胞周期调控、DNA损伤修复、细胞凋亡等相关。实验结果为阐明重离子辐射效应的分子机制提供了有用信息。在进行上述实验研究的同时,学习和掌握了辐射致DNA和细胞损伤的几种检测技术,并对它们的适用性和优缺点进行了评估。更多还原

【Abstract】 Proton and heavy ions have been applied in cancer radiation therapy field, and people have accepted this method increasingly. Under this background, the research on the biological effects and mechanisms of mammalian cell irradiated by high linear energy transfer (LET) ions has become more and more important.This research investigate the biological effects of plasmid DNA and several cells after irradiation by methods of agarose gel electrophoresis along with advanced gel imaging system, colony forming ability experiment along with other traditional cell biology techniques and gene chip as a modern molecular biotechnology. The experimental results of this research can be expected to offer valuable basic data for cancer therapy by heavy ions and protons radiation method, Boron Neutron Capture Therapy (BNCT) as well as the dangerous assessment and protection of human in space or nuclear radiation environment, etc. The experimental procedure and elementary results are as follows:The pUC19 plasmid DNA with or without free radical scavengers in aqueous solution is irradiated by 60Co-γrays, the results are analyzed by gel electrophoresis and Alpha Innotech digital imaging system .The results indicate that free radicals produced byγrays radiation are an important factor for the induction of DNA single strand breaks (SSB) and DNA double strand breaks (DSB). Mannitol, Vitamin C and tea polyphenols are all good scavengers, and mannitol is more effective than Vitamin C under the same condition. Compared with heavy ions, the capacity of scavengers is more remarkable on free radicals induced byγrays, which is more important to evaluate the effect of free radicals in heavy ion radiation.U251 glioma cell and HepG2 liver cancer cell are irradiated by 60Co-γrays andαparticles emitted from 238Puαsource respectively. The results indicate that U251 cell is more sensitive to ionizing radiation than HepG2 cell, andαparticles irradiation inhibit the cell proliferation more effectively thanγrays irradiation. It means thatαparticles have a higher relative biological effectiveness (RBE) thanγrays.L02 normal liver cell is irradiated by 5Gy 80MeV/u 12C6+ ions in radiation terminal of Heavy Ion Research Facility at Lanzhou (HIRFL). 22k human genome oligonucleotide microarray was used to explore the transcriptional profiles of the cells at 6h and 24h post-irradiation. In the 6h post-irradiation cells, the microarrays displayed 37 differentially expressed genes, including 18 up-regulated genes and 19 down-regulated genes. In the 24h post-irradiation cells, the microarrays displayed 269 differentially expressed genes, including 67 up-regulated gene and 202 down-regulated genes. Most of these changed genes were associated with signal transduction,cell cycle control,DNA repair and apoptosis. These observations provided the information of radiation-induced genes for elucidating molecular mechanisms of the diverse biological effects generated by heavy ion irradiation.During the experiment mentioned above, several testing techniques on DNA and cell damage have been learnt and the adaptability,advantages and disadvantages of these methods are evaluated.更多还原