1994 Fiscal Year Final Research Report Summary
Molecular analysis for the etiology of Wilson disease
| Project/Area Number |
05670704
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
Pediatrics
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| Research Institution | Toho University |
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Principal Investigator |
AOKI Tsugutoshi Toho University, 2nd Department of Pediatrics, professor, 医学部, 教授 (50057585)
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| Co-Investigator(Kenkyū-buntansha) |
UCHIYAMA Toshimitu Toho University, Department of Pharmacology, professor, 医学部, 教授 (50057709)
HEMMI Hiromichi Toho University, Laboratry of Molecular Biology, associate professor, 医学部, 助教授 (90165514)
SHIMATAKE Hiroyuki Toho University, Laboratory of Molecular Biology, professor, 医学部, 教授 (40010110)
FUJIOKA Yoshimi Toho University, 2nd Department of Pediatrics, staff, 医学部, 助手 (30256739)
SHIMIZU Norikazu Toho University, 2nd Department of Pediatrics, staff, 医学部, 助手 (60256740)
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| Project Period (FY) |
1993 – 1994
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| Keywords | Wilson disease / Copper metabolism / ceruloplasmin / p-type ATPase / Anti-holoceruloplasmin monoclonal antibody |
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| Research Abstract |
Wilson disease is an autosomal recessive disorder of copper metabolism and has been mapped to the long arm of chromosome 13q14.3. In this disease, copper accumulation in the liver is followed by accumulation in the kidney, cornea and brain tissue. As the feature laboratory findings, serum level of ceruloplasmin, that is a blue multi-copper oxidase, is low in 95% of patients with Wilson disease. In the end of 1993, the candidate gene of Wilson disease has been cloned, and some mutation of this gene from patients with Wilson disease were reported. This gene encodes a copper transporting P-type ATPase. We investigated molecular analysis of this gene for the patients with Wilson disease, and also analyzed relationship for holo and apo-ceruloplsmin of this disease.
We measured the level of holoceruloplasmin and total ceruloplasmin by ELISA method, using antitotal and antiholoceruloplasmin, on sera of the patients with Wilson disease. Serum holoceruloplasmin levels of these patients were remarkably low, and the ratio of holoceruloplasmin for total ceruloplasmin were 30-80%, while that of normal subjects were more than 92%. From these data, we conclude that the etiology of hypoceruloplasminemia of Wilson disease is a defect for copper binding step of ceruloplasmin.
We isolated genomic DNA and total RNA from peripheral blood and liver of patients with Wilson disease and normal subjects. Southern blot analysis and Northern blot analysis were performed using Wilson disease gene cDNA as a probe. Wilson disease patients has no major deletion or insertion, more than several hundred basepaire. Some RFLP patterns were detected, so we speculate that this study is useful to familial analysis of this disease. Northern blot analysis revealed the expression of Wilson disease gene of patient's liver was qualitatively and quantitatively similar as control. This result suggested that the gene mutation of this patient does not influence mRNA expression.
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