1989 Fiscal Year Final Research Report Summary
Developments of the diagnostic methods in heritable connective tissue diseases on the enzymatic and cDNAs levels
| Project/Area Number |
62870042
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| Research Category |
Grant-in-Aid for Developmental Scientific Research
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| Allocation Type | Single-year Grants |
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| Research Field |
Dermatology
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| Research Institution | Medical College of Oita |
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Principal Investigator |
SHINKAI Hiroshi Medical College of Oita, Med. Assist. Prof., 医学部, 助教授 (90030957)
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| Co-Investigator(Kenkyū-buntansha) |
KATAGIRI Kazumoto Medical College of Oita, Med. Assistant, 医学部, 助手 (00204420)
HONDA Tomohito Medical College of Oita, Med. Assistant, 医学部, 助手 (80173665)
MATSUNAGA Etsuji Medical College of Oita, Med. Lecturer, 医学部, 講師 (00145400)
FUJIWARA Sakuhei Medical College of Oita, Med. Lecturer, 医学部, 講師 (90181411)
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| Project Period (FY) |
1987 – 1989
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| Keywords | Collagen / Hydroxyproline / Hydroxylysine / Ehlers-Danlos / Proteodermatan sulfate / HPLC |
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| Research Abstract |
Molecular defects of collagens or proteoglycans including the replacement of amino acid in polypeptide chains and defects of the modified enzymes of their propeptides, have been found in heritable connective tissue diseases. Amino acid analyses using HPLC system were simply revealed the hydroxylysine deficient case in a patient with Ehlers-Danlos syndrome from the newly synthesized collagen of the culture cells. Northern and southern blot analyses using cDNA of collagen alpha 1(I)- and alpha 2(I)-chain, showed depression of alpha 2(I) gene expression from a variant form of Ehlers-Danlos syndrome. The study of collagen structure, in health and disease, frequently requires fractionation and peptide analysis of the collagen chains. In order to micro-analysis of the heterogeneity of newly synthesized collagen from hereditary connective tissue diseases, reverse-phase HPLC system was more useful for the rapid separation of the CNBr peptides from each alpha- chain that was separated by ion exchange chromatography by HPLC. Two dimensional electrophoresis using SDS-polyacrylamid gel (5%) as first dimension and SDS-polyacrylamid gel electrophoresis as second after the treatment of the gel with CNBr was useful for the detection of the molecular heterogeneity of collagen. Dermatan sulfate proteoglycans which associate with collagen fibril, were found several forms in interstitial connective tissues. The defect of dermatan sulfate proteoglycans was found in a case of Ehlers- Danlos syndrome. In order to more clarify the cause of the proteoglycan deficiency, we attempt to cDNA cloning from the culture fibroblasts and its sequences.
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