1997 Fiscal Year Final Research Report Summary
A FUNDAMENTAL RESEARCH FOR GENE THERAPY OF HEMOPHILIA A-EXPRESSION AND RECONSTITUTION OF FACTOR VIII SUBUNITS-
| Project/Area Number |
08670908
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Pediatrics
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| Research Institution | NARA MEDICAL UNIVERSITY |
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Principal Investigator |
TANAKA Ichiro NARA MEDICAL UNIVERISTY,MEDICALDEPARTMENT,ASSISTANT PROFESSOR, 医学部, 講師 (00201616)
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| Co-Investigator(Kenkyū-buntansha) |
NAKA Hiroyuki NARA MEDICAL UNIVERISTY,MEDICALDEPARTMENT,ASSISTANT PROFESSOR, 医学部, 助手 (40281761)
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| Project Period (FY) |
1996 – 1997
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| Keywords | factor VIII / subunit / gene therapy |
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| Research Abstract |
Each of approximately 2kb of cDNA,corresponding to factor VIII heavy and light chain, was individually inserted into the expression vector pNeoIG502. After CHO (Chinese Hamster Ovary) cells were transfected with the constracted vectors, cell supernatants were tested for the presence of factor VIII coagulant activity using both clotting and chromogenic assays, and factor VIII antigen using subunit-specific ELISAs. In the supernatant 0.02-0.04U/ml of light chain subunit was detected, whereas no heacy chain subunit was detected using subunit-specific ELISAs. Factor VIII coagulant activity was also undetectable by both clotting and chromogenic assays. When adenoviral vector was used as the expression vector instead of pNeoIG502,0.1-0.2U/ml of light chain subunit was detected in the supernatants of cells with no detection of heavy chain and coagulant activity of factor VIII.
Further investigations have been attemptted using various vectors, such as retroviral or adeno-associated viral vector as the expression vector, and hepatoma cells (HepG2), fibroblast cells, or myoblast cells as the target cell.
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