| Project/Area Number |
14571677
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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 |
Ophthalmology
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| Research Institution | Saga University |
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Principal Investigator |
KOBAYASHI Hiroshi Saga University, Faculty of Medicine, Ophthalmology, Associate Professor, 医学部, 助教授 (00215360)
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| Co-Investigator(Kenkyū-buntansha) |
KOBAYASHI Kaori Saga University, Faculty of Medicine, Ophthalmology, Assistant Professor, 医学部, 講師 (60325595)
中林 條 佐賀医科大学, 医学部, 助手 (10315202)
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| Project Period (FY) |
2002 – 2003
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| Project Status |
Completed (Fiscal Year 2003)
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| Budget Amount *help |
¥4,000,000 (Direct Cost: ¥4,000,000)
Fiscal Year 2003: ¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 2002: ¥2,300,000 (Direct Cost: ¥2,300,000)
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| Keywords | Retinal pigment epithelium / siRNA / Silencing / pP344 / セリンプロテアーゼインヒビター / 免疫組織化学法 / ウエスタンブロット法 |
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| Research Abstract |
It is appropriate to use non-pathogenic virus with the promoter of pP344 gene to develop the delivery system which provides the specific expression of a target gene in the retinal pigment epithelium. We thought that not only isolation of pP344 promotor but also study of distribution of pP344 product was important. We raised antibodies against PP344 protein to investigate its ocular distribution. lmmunohistochemical study showed. the significant staining in the RPE and the entire neural retina in chick embiyo eyes and mouse eyes. Western blotting analysis revealed the 65 kDa band of cell extract of RPE and neural retina, Therefore, the PP344 protein was secreted to the apical side and be distributed in the neural retina, and may play an important role in the functions of neural retina. We also addressed on the way of the control of gene expression. The use of small interfering RNAs (siRNA) has become a powerful tool to knock down specific gene expression in a wide range of cells. To identify the functional rnechanism.of Mitf (microphthalmia-inducing transcription factor) in differentiation, we used siRNA to specifically suppress Mitf expression in RPE and investigate the expression of MMP 115 (melanosomal matrix,protein 115 kDa) in the transdiffei鋲ntiation process. The expression of MMP 115 was extremely reduced by 75 %. The expression of MMP 115 was extremely reduced and the number of differentiated cell was decreased. Mitf expression in RPE cells was efficiently inhibited using RNA interference technology. Knockdown of Mitf in RPE cells results in their, promotion of transdifferentiation.
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