| Project/Area Number |
05454477
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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 |
Ophthalmology
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| Research Institution | Juntendo University |
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Principal Investigator |
KANAI Atsushi Juntendo Univ., Dept.of Ophthalmol.Professor, 医学部, 教授 (00053059)
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| Co-Investigator(Kenkyū-buntansha) |
SAKUMA Hitoshi Juntendo Univ., Dept.of Ophthalmol.Assistant, 医学部, 助手 (60235207)
FUJIKI Keiko Juntendo Univ., Dept.of Ophthalmol.Assistant Prof., 医学部, 講師
HOTTA Yoshihiro Juntendo Univ., Dept.of Ophthalmol.Assistant Prof., 医学部, 講師 (90173608)
NAKAYASU Kiyoo Juntendo Univ., Dept.of Ophthalmol.Associate Prof., 医学部, 助教授 (10124976)
矢島 寿広 順天堂大学, 医学部, 助手 (50230199)
佐渡 一成 順天堂大学, 医学部, 助手 (70205988)
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| Project Period (FY) |
1993 – 1994
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| Project Status |
Completed (Fiscal Year 1994)
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| Budget Amount *help |
¥4,100,000 (Direct Cost: ¥4,100,000)
Fiscal Year 1994: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1993: ¥3,300,000 (Direct Cost: ¥3,300,000)
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| Keywords | Corneal dystrophy / Gene / Prealbumin / Rabbit / Corneal endothelium / cDNA / Ferritin / FK-506 / rapamycin binding protein / 格子状角膜変性症 / プレアルブミン遺伝子 / SSCP / cDNAライブラリー / プラスマイナススクリーニング / 家兎角膜内皮細胞 / NADH-ユビキノン酸化還元酵素 |
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| Research Abstract |
1. Candidate gene screening in corneal dystrophies
We analyzed the prealbumin gene using SSCP (single strand conformation polymorphism), in 8 unrelated Japanese corneal dystrophies, gelatinous drop-like, lattice, granular, and macular corneal dystrophy patients. The DNA fragments for each exon were amplified by PCR (polymerase chain reaction) and were tested by SSCP method. No mutation was detected in the prealbumin gene.
2. Cloning of the rabbit corneal endothelial cDNA
The rabbit corneal endothelial cDNA library (Yamaguchi et.al., J.Biol.Chem., 1989) was employed. Plus minus screening was performed in 1000 clones usiong corneal and iris total RNA of rabbit as probes. Twenty clones which were positive to the corneal probe and negative to the iris probe were obtained. Regions of inserts of clones were amplified by PCR and directly sequenced. Sequenced data were analyzed using Gene Works and EMBL.There were several clones that showed high homology with each knowen cDNA ; B22 subunit of bovine mitochondrial NADH-ubiquinone oxidoreductase, human and cattle FK-506/rapamycin binding protein 25 (FKBP25), human thrombospondin 2 and rabbit ferritin. 5 unknown clones that showed no homology with any previous reported cDNA were also isolated. The rabbit FKBP25 cDNA clone has the entire coding sequence except an initiation codon and 3' untranslated sequence until poly A.RT-PCR analysis showed high expression of FKBP25 mRNA in cornea. Since the calcium release channel of endoplasmic reticulum is modulated by FKBP and Ca^<2+> plays a important role in pump function of corneal endothelium. FKBP may play an great role in the pomp function in corneal endothelium. Although further study is required, these clones may become the candidate gene for the corneal dystrophies. Furthermore, some of them seem to be important for corneal research.
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