The study of cauces of mitochondrial cytopathy

大多数线粒体酶蛋白在细胞质中作为前体酶蛋白合成，随后进入线粒体形成成熟的酶蛋白。该输入系统的缺陷可能是由于线粒体功能紊乱而引起具有严重临床症状的线粒体细胞病。近年来，我们发现了一例含丙酮酸脱氢酶（PDH）复合物的线粒体多酶缺陷患者。在本项目中，为了确定该患者的主要缺陷，我们使用细胞标记和脉冲追踪方法检测了PDH前体蛋白（pEla和pElb）进入患者成纤维细胞线粒体的输入系统。患者的成纤维细胞中pEla和pElb进入线粒体的比率分别下降到正常对照成纤维细胞的40%和63%左右。这些结果表明，该患者线粒体中Ela Elb的输入系统存在缺陷。由于患者不仅存在PDH复合物缺陷，其他线粒体酶也存在缺陷，且患者体内pEla和pElb的cdna未发生突变，因此该患者的主要缺陷是线粒体酶蛋白进入线粒体的进口系统。在另一项研究中，我们在女性PDH缺乏症患者的PDH亚基基因中发现了一个新的18bp插入突变，PDH缺乏症是线粒体细胞病变的常见原因，并且该专利培养的细胞中突变基因的表达存在广泛差异。提示女性PDH缺乏症仅用酶学诊断有可能被误诊。因此，有必要开发新的诊断方法，如DNA分析。

Most of mitochondrial enzyme proteins are synthesized in the cytoplasm as precursor enzyme proteins and are subsequently imported into mitochondria to form mature enzyme proteins. The defect in this import system might be cause the mitochondrial cytopathy with sever clinical symptoms due to disturbance of mitochondrial functions. Recently we found the patient with defect of multiple mitochondrial enzymes containing pyruvate dehydrogenase (PDH) complex. In this project, to determine the primary defect in this patient we examined the import system of precursor proteins of PDH (pEla and pElb) into mitochondria in fibroblasts from the patient using cell-labeling and pulse-chase methods. The rates of import of pEla and pElb into mitochondria in fibroblasts from the patient decreased to about 40% and 63% of those in normal control fibroblasts, respectively. These results showed the defect of import system of Ela na Elb into mitochondria in this patient. Since the patient had not only the defect of PDH complex but also the defect of other mitochondrial enzymes and cDNAs for pEla and pElb in the patient had no mutation, the primary defect in this patient is in the import system of mitochondrila enzyme proteins into mitochondria.In anotheer study, we found a new 18bp insertion mutation in the gene for a subunit of PDH in afemale patient with PDH deficiency, the common cause of mitochondrial cytopathy, and the wide variation in the expression of mutant gene in cultcred cells from this patent. These results showed that female patients with PDH deficiency might be misdiagnosed only with enzymological diagnosis. Therefore, it is necessry to develop the new diagnosis method of PDH deficiency, such as DNA analysis.