MOLECULAR AND BIOCHEMICAL ANALYSIS OF THE CARNITINE CYCLE DISORDERS AS A POSSIBLE CAUSE OF AN ACUTE ENCEPHALOPATHY AND REYE-LIKE SYNDROME

肉碱循环紊乱作为急性脑病和雷氏综合征可能病因的分子和生物化学分析

基本信息

批准号：
11670739
负责人：
KANAZAWA Masaki
金额：
$ 2.37万
依托单位：
CHIBA UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2001
项目状态：
已结题

项目摘要

We collected samples (blood, urine, lymphocytes and fibroblasts) from Japanese patients with recurrent hypoketotic hypoglycemia, acute encephalopathies or Reye-like syndrome, and biochemicaly, enzymaticaly, and genetically analyzed them with parental permission, to confirm these patients were affected with the carnitine cycle disorders.A 3 years old boy presented with typical Reye-like syndrome with unconsciousness, hyperammonemia, hypoglycemia, and low carnitine level, and diagnosed deficiency of carnitine transporter. This was confirmed by enzyme analysis in fibroblast cultures. A mutation analysis of the SLC22A5 gene is under investigationWe analyzed cDNA and genomic DNA of CPT1A in four patients with liver carnitine palmitoyltransferase I (CPT1-L) deficiency presenting with Reye-like syndrome. Analysis of the gene revealed that patient 1 was compound heterozygous for 1425G>A (W475X) and 1494T>G (Y498X) ; patient 2 was compound heterozygous for 96T>G (Y32X) and 1079A>G (E360G) ; pat … More ient 3 was homozygous for 281+1insT ; patient 4 carried 2027-2028+2delAAGT in the paternal allele. Six novel mutations have been identified in four patients, confirming that mutations in CPT1A are responsible for CPT1-L deficiency presenting with Reye-like syndrome. One of these mutations is a missense mutation, E360G. The other is a splicing mutation, 2027-2028+2delAAGT, which causes aberrant splicing transcripts, 1876-2028del, 2027-2028insGTCTCTTCCACTTCTTCC and 2026-2028del. These three aberrant transcripts keep reading flame. Expression study using SV40 transformed fibroblasts was performed to investigate the consequences of these two mutations on enzyme activity and protein level. Molecular analysis in this study revealed that these two mutations were the disease-causing mutations.Fibroblasts obtained from a patient with carnitine acylcarnitine carrier (CAC) deficiency were purchased from the National Institute of General Medical Sciences (NIGMS). We have identified two novel mutations of this patient The first, a deletion mutation (146 del T), leads to premature termination and results in a very immature CAC protein. The second, a splicing mutation, (261-10T>G) results in either skipping exons 3 and 4, or of exon 3 alone, and leads to truncation of protein. These data were published in Journal of Human Genetics (2000).These results were contributed to a therapeutic approach and an improvement of prognosis of the patients with the carnitine cycle disorders. Less

我们从日本复发性低血糖，急性脑病或雷耶综合症或生物化学的疾病中，与这些典型的典型分析的典型分析的日本患者相比，我们收集了日本患有复发性低血糖，急性脑病或雷耶综合征或生物化学的样品（血液，尿液，淋巴细胞和成纤维细胞）的样品（急性脑病或雷耶综合症，生物化学综合症，并与养生的典型分析，都与养生的典型分析。 Reye样综合征，无意识，高症，低血糖症和低肉碱水平以及肉碱转运蛋白的诊断缺乏。通过成纤维细胞培养物中的酶分析证实了这一点。正在研究SLC22A5基因的突变分析我们分析了四名患有雷耶样综合征的肝脏肉碱棕榈酰转移酶I（CPT1-L）缺乏症的cpt1a的cPT1A和基因组DNA。该基因的分析表明，患者1的1425G> A（W475X）和1494T> G（Y498X）是复合杂合的；患者2对96T> g（y32x）和1079a> g（e360g）的复合杂合子; PAT…更多的Ient 3是281+1inst的纯合子；患者4在父亲等位基因中携带2027-2028+2Delaagt。在四名患者中已经确定了六个新型突变，证实CPT1A中的突变是伴有Reye样综合征的CPT1-L缺陷。这些突变之一是错义突变，e360g。另一个是剪接突变，2027-2028+2delaagt，导致异常剪接转录本，1876-2028del，2027-2028insgtcttcttccatttcttcttcttcc和2026-2028del。这三个异常笔录不断阅读火焰。使用SV40转化的成纤维细胞进行了表达研究，以研究这两个突变对酶活性和蛋白质水平的后果。这项研究中的分子分析表明，这两个突变是引起疾病的突变。从肉碱酰基肉碱载体（CAC）缺乏症患者获得的纤维细胞购自美国国家一般医学科学研究所（NIGMS）。我们已经确定了该患者的两个新型突变，即缺失突变（146 del t），导致过早终止，并导致非常不成熟的CAC蛋白。第二个是剪接突变，（261-10t> g）导致跳过外显子3和4，或单独使用外显子3，并导致蛋白质的截断。这些数据发表在《人类遗传学杂志》（2000年）中。这些结果有助于一种治疗方法，并改善了肉碱循环疾病患者的预后。较少的