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

我们从患有复发性低酮性低血糖、急性脑病或雷氏样综合征的日本患者中采集了样本（血液、尿液、淋巴细胞和成纤维细胞），并在父母许可的情况下对其进行生化、酶学和基因分析，以确认这些患者受到肉碱的影响循环障碍。3岁男孩综合征表现为典型的雷氏综合征，伴有意识不清、高氨血症、低血糖和低肉碱水平，以及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 4 名患有 Reye 的肝肉碱棕榈酰转移酶 I (CPT1-L) 缺乏症患者的 CPT1A cDNA 和基因组 DNA。基因分析显示，患者 1 为 1425G>A (W475X) 复合杂合子，并且1494T>G (Y498X) ；患者 2 为 96T>G (Y32X) 和 1079A>G (E360G) 复合杂合；患者 3 为 281+1insT 纯合；患者 4 的父系携带 2027-2028+2delAAGT在四名患者中发现了六种新的突变，证实了该突变。 CPT1A 中的 CPT1-L 缺陷导致雷耶样综合征，其中一种突变是错义突变，E360G，另一种是剪接突变，2027-2028+2delAAGT，它会导致异常剪接转录本，1876-2028del。 2027-2028insGTCTCTTCCACTTCTTCC 和2026-2028del.使用 SV40 转化的成纤维细胞进行表达研究，以研究这两种突变对酶活性和蛋白质水平的影响，表明这两种突变是致病原因。我们已鉴定从肉碱酰基肉碱载体 (CAC) 缺乏症患者获得的成纤维细胞购自美国国立普通医学科学研究所 (NIGMS)。该患者的两个新突变第一个是缺失突变 (146 del T)，导致过早终止并产生非常不成熟的 CAC 蛋白，第二个是剪接突变 (261-10T>G) 导致外显子跳跃。 3 和 4 或单独的外显子 3，并导致蛋白质截短。这些数据发表在《人类遗传学杂志》(2000) 上。这些结果有助于治疗方法和改进。肉毒碱循环障碍患者的预后较差。