利用细胞衰老模型探讨线粒体与衰老的关系

发布时间：2018-01-21 10:14

本文关键词： 线粒体线粒体DNA 人衰老 W1-38细胞系 PCR γ射线膜电位氧化还原酶　出处：《第二军医大学》2005年硕士论文　论文类型：学位论文

【摘要】：线粒体作为真核细胞中一种重要而独特的细胞器,其性状改变与人的衰老和疾病密切相关,甚至被称为衰老的生物钟。自1989年Linnane等提出线粒体衰老假说以来,人们越来越关注线粒体与衰老的关系。因线粒体一些特有的特点,如生命周期有限,具有高的更新率;形态、大小、数量和分布,常因细胞种类不同而异;线粒体DNA(mtDNA)与核DNA不同,是多拷贝的且具有异质性等,到目前为止,虽对衰老中线粒体的改变有一些研究报道,但结果并不一致,缺乏较系统的研究。关于人衰老中线粒体的一些变化并无定论,线粒体衰退与机体的衰老二者的因果关系也有待于进一步探讨。机体的衰老来源于细胞的衰老,对二倍体细胞的研究有助于衰老机理的探讨。通过对二倍体细胞衰老中线粒体变化的研究,有助于探讨线粒体与衰老的关系,进一步弄清线粒体在衰老中的作用。这些研究将对衰老指标的测定,抗衰老药物研究及措施的评估有重要意义。本实验主要通过建立细胞衰老模型,从二倍体细胞衰老过程中量与功能变化相结合来探讨线粒体与衰老的关系。一.人胚肺二倍体细胞W1-38的培养近年来应用人胚肺二倍体细胞W1-38作为衰老研究的体外模型,己得到了国内外研究工作者的肯定。细胞从20代传至约50代;定期冻存;隔期拍照记录生长状态;计数板计数;MTT测定细胞活力。观察发现,年轻细胞平均两天呈致密单层,细胞长梭形,排列成束状,折光度好,立体感强;每瓶细胞数为6～8×10~5;衰老细胞形成单层时间明显延长,细胞圆缩蜕变,可呈菱形、三角形,折光度下降;每瓶细胞数为4～6×10~5。晚期细胞活力明显低于早期。二.衰老中线粒体质和量改变的研究线粒体形态因外界环境和功能改变而异,线粒体形态变化可作为衰老研究的参考之一。收集固定细胞,制成切片,电镜观察线粒体三维结构和内部结构。线粒体的数量在不同细胞中差异很大,同时与细胞的功能状态是相关的。
[Abstract]:Mitochondria, as an important and unique organelle in eukaryotic cells, are closely related to human aging and disease. Since Linnane et al put forward the hypothesis of mitochondrial senescence in 1989, people have paid more and more attention to the relationship between mitochondria and aging. Because of some unique characteristics of mitochondria, such as limited life cycle, has a high rate of renewal; The shape, size, quantity and distribution of the cells often vary from cell to cell. Mitochondrial DNA (mtDNA), unlike nuclear DNA, is multicopy and heterogeneous. Up to now, there have been some studies on mitochondrial changes in aging, but the results are not consistent. Lack of systematic research. There is no conclusion about some changes of mitochondria in human aging. The causal relationship between mitochondrial decline and aging of organism needs to be further discussed. The senescence of organism comes from cell senescence. The study of diploid cells is helpful to probe into the mechanism of senescence and the relationship between mitochondria and senescence by studying the changes of mitochondria in diploid cells. These studies will be of great significance to the determination of aging indexes, the study of anti-aging drugs and the evaluation of measures. In this experiment, the relationship between mitochondria and senescence was studied by establishing the model of cell senescence and combining the changes of quantity and function during the aging of diploid cells. 1. Culture of human embryonic lung diploid cells W1-38 In recent years, the application of human embryonic lung diploid cell W1-38 as an in vitro model of aging research has been confirmed by researchers at home and abroad. Regular freezing; The growth state was recorded by taking pictures at intervals. Counting board; Cell viability was measured by MTT. The results showed that the young cells were compact monolayer on average for two days with long fusiform cells arranged in bundles good diopter and strong stereosensitivity. The number of cells per bottle was 6 ~ 8 脳 10 ~ (5); The monolayer formation time of senescent cells was obviously prolonged, the cells were round and degenerated, and could be rhomboid, triangular, and diopter decreased. The number of cells per bottle was 4 ~ 6 脳 10 ~ (5). The activity of late stage cells was significantly lower than that of early stage. II. Studies on changes of mitochondrial quality and quantity in aging The morphology of mitochondria varies with the changes of environment and function. The changes of mitochondrial morphology can be used as a reference for the study of aging. Fixed cells are collected and made into sections. Electron microscope was used to observe the three-dimensional and internal structure of mitochondria. The number of mitochondria varied greatly in different cells and was related to the functional state of the cells.
【学位授予单位】：第二军医大学
【学位级别】：硕士
【学位授予年份】：2005
【分类号】：R33

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