1992 Fiscal Year Final Research Report Summary
Molecular Pathogenesis of a Dominantly Inherited Disorder-Analysis by Using a Mouse Model of Disorder
| Project/Area Number |
02454494
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
Human genetics
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| Research Institution | KUMAMOTO UNIVERSITY |
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Principal Investigator |
MAEDA Shuichiro Kumamoto Univ.School of Medicine, Dept.of Biochemistry, Assoc.Prof., 医学部, 助教授 (10117244)
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| Co-Investigator(Kenkyū-buntansha) |
SETOYAMA Chiaki Kumamoto Univ.School of Medicine, Dept.of Biochemistry, Lecturer, 医学部, 講師 (60040250)
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| Project Period (FY) |
1990 – 1992
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| Keywords | Amyloidosis / Serum amyloid P component / Mouse model of disorder / Transgenic mouse / Transthyretin / Gene targeting / Familial amyloidotic polyneuropathy / Dominantly inherited disorder |
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| Research Abstract |
(1) We examined whether sustained high serum levels of serum amyloid P component (SAP) induced by repeated intraperitoneal injections of E.coli lipopolysaccharide (LPS) enhance the amyloid deposition in the transgenic mouse model of familial amyloidotic polyneuropathy (FAP). No significant difference was detected in the onset, progression, and tissue distribution of amyloid deposition between the LPS-stimulated and unstimulated transgenic mice.
(2) We constructed a sequence replacement vector to introduce an inactivating neo^r marker into the mouse genomic transthyretin (ttr) gene by homologous recombination. The targeting vector consisted of a 5.9 kb DNA fragment derived from the mouse ttr genomic locus and the 1.1 kb neo^r gene from pMC1Neo. A counter-selectable HSV-tk gene was placed at the end of the long region of homology. This vector was electroporated into cultured mouse embryonic stem (ES) cells, which were subsequently selected with G418 and gancyclovir. Among the 132 colonies obtained, 6 homologous recombinants were identified by PCR and Southern blotting.
(3) These ES cells were used to generate chimeric mice by injection into blastocysts. One of these chimeras has shown germline transmission of the disrupted ttr gene. Heterozygous mice were crossed to try to generate homozygotes. Homozygous TTR-deficient mice were found at the expected Mendelian frequencies and appeared to be phenotypically normal. However, levels of serum retinol, retinol-binding protein, and thyroid hormone are significantly depressed in the mutant mice. We are now in the process of establishing transgenic mouse lines carrying the human mutant ttr gene which is responsible for FAP in the homozygous mutant background.
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Research Products
(20 results)
