脑损伤基因差异表达与损伤时间推断的实验研究

【作者】 李廷富；

【导师】 万立华；

【作者基本信息】 重庆医科大学 ， 临床检验诊断学， 2004， 博士

【摘要】 目的:研究创伤性脑损伤早期基因表达谱与正常脑组织基因表达谱的差异,以期阐明脑损伤后早期基因表达的改变规律,阐明脑损伤发生发展的分子机制，从而为临床治疗提供帮助,同时为法医损伤时间推断研究寻找标志物提供帮助。方法：以大鼠自由落体损伤模型为对象，从损伤区脑组织和假手术对照组脑组织分别提取mRNA,经反转录成cDNA后与含有4096个随机基因的基因表达谱芯片杂交，杂交后的芯片经扫描仪扫描，并用GenePix3.0软件分析结果。结果:发现有124个差异表达基因或表达序列标签(Expression Sequence Tags ESTs);其中有46个基因和26个EST表达下调; 28个基因和24个EST表达上调;在这些表达有差异的基因中,有涉及细胞内信号传导、神经递质释放、参与炎症的蛋白、离子通道及其受体蛋白和参与炎症反应的蛋白等被发现。结论：创伤性脑损伤的发生发展涉及多个基因的改变；研究一个或少数几个基因很难解释其损伤后分子变化机制；基因芯片是研究颅脑损伤这种多基因改变、多因素作用的理想工具。更多还原

【Abstract】 Objective:To study the expression levels of large number of genes and to identify genes not previously implicated in rat after traumatic brain injury,which may provide valuable information on therapies and may be helpful in search of forensic markers of vitality.Methods:The model of traumatic brain injury originally <WP=13>described by Feeney was employed with modification.Wistar rats were randomly divided into cortical impact injury group and shame operated control group.Each group comprised 12 Wistar rats. At 5 hours following cortical impact injury, animals were anaesthetised by intraperioneal injection of 4% chloral hydrate and sacrificed by decapitation .Then the brain tissue were dissected and stored in the liquid nitrogen until required for RNA isolation. The total RNA of tissues were extracted and the probes were prepared by reverse transcriptase reaction(RT)and then hybridized with cDNA microarray.Results:Of 4096 randomly selected arrayed genes or expressed sequence tags from a rat cDNA library,124 genes or expressed sequence tags were found to be differently expressed at 5 hours following cortical impact injury(28 genes and 24 ESTs were increased, and 46 genes and 26 ESTs were decreased);Genes involved in cell homeostasis such as Secretory leukocyte protease inhibitor, transthyretin and inflammatory response such as phospholipase A2,interliukin-1 were primarily up-regulated while those encoding mitochondrial enzymes(such as ATP synthase,ATP citrate lyase),metobolic molecules, calcium signaling protein ,and cellular structure protein(such as tublin) were predominantly down-regulated; Down-regulated <WP=14>genes were also seen in neutrophic factor/receptors(Ciliary neutrophic factor, Glial cell line-derived neutrophic factor ); Partly channels and transporters were down-regulated whereas partly channels and transporters were up-regulated at same time.To our best knowledge,A number of genes were found which have not been previously report in traumatic brain injury.Conclusions:The geneomic response to traumatic brain injury is complex and a great number of genes involved in this pathophysiological course. The molecular mechanism of traumatic brain injury could not be fully disclosed by investigating only one changes of a gene or a few genes.The microarray methods is an ideal tools for researching multi-genes involved diseases.更多还原