MECHANISMS OF EXPRESSION OF DELETED MITOCHONDRIAL DNA FROM CULTURE CELLS

我们建立的方法用于肌肉、皮肤和淋巴细胞的冷冻保存和后续培养，可以在体外完成异常线粒体DNA的维持和细胞表型特征的保持。培养的细胞包括肌肉、皮肤成纤维细胞和淋巴细胞，来自200例临床诊断为线粒体异常的患者的2000多个样本。这些细胞用于分析异常线粒体DNA的基因表达。然后，我们尝试开发了这些细胞信息的数据库软件，以备将来使用。慢性进行性眼外肌麻痹(CPEO)是线粒体脑肌病的一个亚型，线粒体DNA大量缺失的突变常见于CPEO患者。为了排除核基因组参与CPEO患者线粒体功能障碍的表达，我们将CPEO患者的mtDNA导入无mtDNA的克隆性HeLa细胞，并分离出环状克隆。我们试图通过用溴化乙锭灭活线粒体DNA来建立无线粒体的细胞系，以研究观察到的异常是否来自线粒体基因的核基因。

Our method developed for cryopreservation and subsequent culture of muscle, skin and lymphocytes could accomplish the maintenance of abnormal mitochondria DNA and the cellular phenotypic characteristics in vitro. The cultured cells including muscle, skin fibro-blast and lymphocytes were more than two thousand samples from two hundred patients suspected with mitochondrail abnormalities by the clinical diagnosis. These cells were used for the analysis of the gene expression of abnormal mitochodrial DNA. We then attempted to develop the software for database about the information of these cells for future use.2. Mutant mitochondrial DNA with large-scale deletions have been frequently observed in patients with chronic progressive external opthalmolplegia(CPEO), a sub group of the mitochondrial encephalomyopathies. To exclude involvement of the nuclear genome in expression of the mitochondrial dysfunction characteristic of CPEO, we introduced the mtDNA of CPEO patient into clonal mtDNA-less HeLa cells, and isolated cybrid clones.3. We attempted to establish cell lines without mitochodria by inactivating mitochondrial DNA with the treatment of ethidium bromide to investigate whether the observed abnormality comes from nuclear genes of mitochondrial genes.