MOLECULAR BASIS OF MITOCHONDRIAL RNA PROCESSING SYSTEM IN DEVELOPMENTAL TISSUES AND IN MITOCHONDRIAL MYOPATHY.

1)为了了解RNA19在正常人体组织中的稳定水平以及在MELAS患者中的实际水平，我们分析了8名正常人各种组织的总RNA，以及4例MELAS-3243点突变患者的肌肉活检组织的总RNA。正常组织包括骨骼肌、脑、心、肾、肝、子宫和脾。用rTth逆转录酶聚合酶链式反应-限制性片段长度多态性方法分析肌肉组织RNA中RNA19组分的点突变百分率。我们观察到，所有4名MELAS患者(是对照组的5-8倍)和1名患有糖原沉积病的患者(是对照组的3倍)的肌肉中RNA19水平显著增加。患者肌肉中RNA19的水平在总ND1信号的20%到35%之间，显著高于对照肌肉(不到总NDI信号的4%)，这与他们有丝分裂…中分析的突变百分比很好地一致更多来自肌肉的线粒体DNA。在肌肉、心脏、脑、肾、肝、脾和子宫等组织中也可识别RNA19，分别占其ND1信号总量的3.5%、11.6%、13.6%、8.4%、4.2%、1.7%和6.5%。在MELAS肌肉中观察到的这种RNA 19的积聚似乎是tRNALeu(UUR)突变所特有的，并与MELAS的发病机制有关。2)研究了5例在线粒体DNA(MtDNA)中存在A3243G突变但具有不同临床表型的无关患者在单个肌肉纤维水平上的突变百分比。1例临床和病理确诊为Leigh综合征(LS)，2例表现为线粒体肌病、脑病、乳酸酸中毒和卒中样发作(MELAS)，1例表现为进行性眼外肌麻痹(PEO)，1例表现为线粒体糖尿病(MDM)。LS患者肌肉的突变量较大(92%)，即使在非粗糙的红纤维(RRF)中也有80%以上的突变，且RRF的比例最高。MELAS患者的RRF突变水平和比例都较低，这两个参数在PEO和MDM患者中甚至更低。这些结果证实了这样一个概念，即不同细胞和组织之间突变载量和突变的空间分布的差异是疾病表型表达差异的原因。较少

1)In order to know the steady-state levels of RNA 19 in normal human tissues and the actual levels of it in MELAS patients, we analyzed the total RNA from various human tissues from 8 normal individuals, and biopsied muscles from 4 MELAS patients having MELAS-3243 point mutation. Normal tissues include skeletal muscle, brain, heart, kidney, liver, uterus, and spleen. We also analyzed the percentage of point mutation in RNA 19 fraction in RNAs from muscles, using rTth reverse transcriptase PCR-RFLP methods. We observed significantly increased levels of RNA 19 in muscles from all 4 MELAS patients (5 - 8 times more than that of the control) and from one patient having glycogen storage disease (3 times more than that of the control). The level of RNA 19 in the patient muscles ranged from 20 to 35% of total ND 1 signal is significantly higher than that of the control muscle (less than 4% of total ND I signal), which is well harmonized with the percentage of mutation analysed in their mitoch … More ondrial DNA from muscles. The RNA 19 is also recognized in various human tissues including muscle, heart, brain, kidney, liver, spleen and uterus, ranging 3.5%, 11.6%, 13 6%, 8.4%, 4.2%, 1.7% and 6.5% of their total ND 1 signal, respectively. This accumulation of RNA 19 observed in MELAS-muscle seems to be specific for the tRNALeu(UUR) mutation and related to the pathogenetic mechanism of MELAS.2)Five unrelated patients harboring the A3243G mutation in the mitochondrial DNA (mtDNA) but presenting with different clinical phenotype were studied for their percentage of mutation at the single muscle fiber levels. One patient had a clinically and pathologically defined Leigh syndrome (LS), two showed mitochondrial myopathy, encephalopathy, lactic acidosis and stroke like episodes (MELAS), another showed progressive external ophthalmoplegia (PEO), and the other showed mitochondrial diabetes mellitus (MDM). The mutation load was greater in the muscle from the patient with LS (92%), who showed more than 80% even in the non-ragged red fibers (RRF) and also presented the highest proportion of RRF.The patients with MELAS had lower mutation levels as well as lower proportion of RRF, and these two parameters were even lower in the PEO and MDM patients. These results confirm the concept that differences in the mutation load and in the spatial distribution of the mutation among different cells and tissues are responsible for the differences in phenotypical expression of the disease. Less