YAMAMOTO Shigenori NATIONAL SHIMOSHIZU HOSPITAL, DEPARTMENT OF CLINICAL RESEARCH, CHIEF OF LABORATORY, 臨床研究部, 生化学研究室長 (30241970)
TERAI Masaru GRADUATE SCHOOL OF MEDICINE, CHIBA UNIVERSITY, DEPARTMENT OF PEDIATRICS, ASSISTANT PROFFESOR, 医学部・附属病院, 講師 (80207472)
金澤 正樹 国立療養所下志津病院, 臨床研究部, 生化学研究室室長代理
|Budget Amount *help
¥3,700,000 (Direct Cost: ¥3,700,000)
Fiscal Year 2001: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 2000: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 1999: ¥1,500,000 (Direct Cost: ¥1,500,000)
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 investigation
We 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
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