早衰小鼠P8（SAMP8）额叶脑衰老相关人鼠同源基因筛选及功能研究

发布时间：2018-06-03 23:45

本文选题：基因芯片 + 脑衰老　；参考：《昆明医学院》2010年博士论文

【摘要】： 随着世界各国人口老龄化进程的到来,研究衰老的机制,进一步防治衰老相关的疾病已逐渐成为当今医学科学研究新的热点。为了更深入地进行脑衰老及相关疾病发生发展分子机制的研究,我们利用目前公认的脑衰老哺乳动物模型——SAMP8(senescence-accelerated mice prone 8,快速老化小鼠-8)——作为研究对象,利用微点阵技术(Microarray/Gene Chip/基因芯片),对12月龄与4月龄SAMP8鼠额叶基因的转录水平进行了对比研究。 Microarray是将指大量寡核苷酸分子固定于支持物上,然后与标记的样品进行杂交,通过检测杂交信号的强弱进而判断样品中靶分子数量的一项新兴生物技术。1995年,Stanford大学的P Brown实验室发明了第一块以玻璃为载体的基因微点阵芯片,标志着基因芯片技术进入了广泛研究和应用的时期。基因芯片被广泛应用于高通量基因表达水平检测、基因诊断、药物筛选及个性化给药、新基因发现等领域,成为21世纪生物技术发展的重要特征。生物信息学(Bioinformatics)是20世纪80年代末随着人类基因组计划(human genome project,HGP)的启动而兴起的一门新的边缘学科和技术,是分子生物学与计算机科学的交叉。其任务之一为发展有效的信息分析工具,构建适合于基因组研究的数据库,搜集、管理和使用人类基因组和模式生物基因组的巨量信息。迄今已发展完善了GenBank核酸序列、SWISS-PROT蛋白质序列和PDB生物大分子结构等著名的数据库,开发了如Blast、NEBcutter、Primer-3等上百种生物信息分析软件。生物信息学就如同一个向导,为科研工作者提供了一个崭新的工具,帮助科研人员从纷繁复杂生物信息中找寻真理。同源基因通常指不同物种中起源于同一祖先的基因,在遗传学中,同源这一概念主要是指序列同源。直向同源序列(Orthologs)被认为在不同的物种中具有相近甚至相同的功能、相似的调控途径,扮演相似甚至相同的角色,而且,绝大多数核心生物功能就是由相当数量的直向同源基因所承担。因此,研究模式生物中与人同源基因的功能对揭示人体各种生理过程及疾病发生发展分子机制具有重要意义。本研究应用基因芯片技术检测12月龄和4月龄SAMP8鼠额叶基因表达谱,筛选到65个明显上调基因和632个明显下调基因。上调基因多为促炎症、应激反应相关基因;而下调基因则多为突触可塑性、囊泡运输、线粒体功能、蛋白质和DNA修复、神经激素等相关基因。进一步经生物信息学分析及RT-PCR、WesternBlot验证,证实了4个基因——Gnaq(Guanine Nucleotide Binding Protein-alpha qPolypeptide)、KIF1b(Kinesin Family Member 1b)、Sort1(sortilin-1)、Sst(somatostatin)——是脑衰老相关人鼠同源基因。这4个基因随衰老呈明显降低的mRNA转录水平和蛋白表达水平。这些发现将有助于更好地在模型动物中研究脑衰老及相关疾病发生的分子机制。基于上述发现,我们进一步研究抗脑衰老药物对脑衰老相关人鼠同源基因表达的影响及这些基因在脑衰老过程中的功能。首先我们研究了天麻素对SAMP8鼠Sst表达水平及相关衰老表征的影响。结果显示天麻素能明显上调SAMP8鼠额叶内Sst的转录水平和蛋白表达水平,并明显改善SAMP8鼠的学习记忆能力和其它衰老相关表征。这些结果提示天麻素治疗可能通过增进额叶Sst的转录和表达水平以延缓SAMP8鼠的脑衰老进程,各种原因所致的Sst表达水平降低可能是导致脑衰老及相关疾病的分子机制之一。Gnaq、KIF1b及Sort1的功能研究正在进行之中。
[Abstract]:With the coming of the aging process of population in the world, the study of the mechanism of aging and the further prevention and control of aging related diseases have gradually become a new hot spot in the research of medical science. In order to further study the molecular mechanism of brain senescence and related diseases, we use the recognized model of brain senescence mammal. - SAMP8 (senescence-accelerated mice prone 8, fast aging mouse -8) - as an object of study, using microarray technology (Microarray/Gene Chip/ gene chip), the transcriptional level of the frontal lobe genes of 12 month old and 4 month old SAMP8 mice was compared.
Microarray is a new biotechnology that is used to immobilize a large number of oligonucleotide molecules on the support and then hybridize with the labeled samples to determine the number of target molecules in the sample by detecting the strength of the hybridization signals in.1995. The P Brown Laboratory of the University of Stanford invented the first gene microdot matrix with glass as the carrier. Gene chip technology has entered a period of extensive research and application. Gene chip has been widely used in high throughput gene expression level detection, gene diagnosis, drug screening and individualization, new gene discovery and so on. It has become an important feature of the development of biotechnology in twenty-first Century.
Bioinformatics (Bioinformatics) is a new frontier discipline and technology that began with the start of the human genome project (HGP) at the end of the 1980s. It is the intersection of molecular biology and computer science. One of its tasks is to develop effective information analysis tools to build data suitable for genome research. Library, collecting, collecting, managing and using the huge information of the human genome and model biological genome. So far, the GenBank nucleic acid sequence, the SWISS-PROT protein sequence and the PDB biological macromolecular structure have been developed, and a hundred kinds of bioinformatics analysis software, such as Blast, NEBcutter, Primer-3, etc. have been developed. A guide provides a new tool for researchers to help researchers find truth from complex biological information.
The homologous gene usually refers to the gene originating from the same ancestor in different species. In genetics, the concept of homologous is mainly the sequence homology. The direct homologous sequence (Orthologs) is considered to have similar or even identical functions in different species, similar to the regulatory pathway, acting similar or even the same role, and the vast majority of the nuclei. The biological function of the heart is assumed by a considerable number of direct homologous genes. Therefore, it is of great significance to study the function of the homologous genes in model organisms to reveal the physiological processes of the human body and the molecular mechanism of the development of the disease.
In this study, gene chip technology was used to detect the gene expression profiles in the frontal lobe of 12 month old and 4 month old SAMP8 mice, and 65 obviously up-regulated genes and 632 down regulated genes were screened. The up regulation was mostly inflammatory and stress related genes, while the down regulated genes were mostly synaptic plasticity, vesicle transport, mitochondrial function, protein and DNA repair, and nerve repair. 4 genes, Gnaq (Guanine Nucleotide Binding Protein-alpha qPolypeptide), KIF1b (Kinesin Family Member), were confirmed by bioinformatics analysis and RT-PCR, WesternBlot verification. These genes are the homologous genes of brain senescence related human mice. These 4 genes are associated with decline. The mRNA transcriptional level and protein expression levels are obviously reduced, and these findings will help to better study the molecular mechanisms of brain senescence and related diseases in model animals.
Based on the above findings, we further studied the effects of anti brain aging drugs on the expression of homologous genes in brain senescence related human mice and the functions of these genes in the process of brain senescence. First, we studied the effect of gastrodin on the expression level of Sst and related senescence in SAMP8 rats. The results showed that gastrodin could significantly increase the Sst in the frontal lobe of SAMP8 rats. The transcriptional level and protein expression level, and obviously improve the learning and memory ability of SAMP8 mice and other aging related characterization. These results suggest that gastrodin therapy may delay the brain aging process in SAMP8 rats by enhancing the transcription and expression level of the frontal Sst, and the decrease of Sst expression caused by various causes may lead to brain senescence and One of the molecular mechanisms of related diseases is the functional study of.Gnaq, KIF1b and Sort1.
【学位授予单位】：昆明医学院
【学位级别】：博士
【学位授予年份】：2010
【分类号】：R339.3

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