| Project/Area Number |
11470172
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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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 | Kumamoto University |
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Principal Investigator |
MIIKE Teruhisa Kumamoto University, Child Development, professor, 医学部, 教授 (90040617)
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| Co-Investigator(Kenkyū-buntansha) |
KIMURA Shigemi Kumamoto University, Child Development, Assistant professor, 医学部附属病院, 助手 (60284767)
藤井 績 熊本大学, 医学部・附属病院, 医員
小川 雅克 熊本大学, 医学部・附属病院, 医員
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| Project Period (FY) |
1999 – 2002
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| Project Status |
Completed (Fiscal Year 2002)
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| Budget Amount *help |
¥13,200,000 (Direct Cost: ¥13,200,000)
Fiscal Year 2002: ¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 2001: ¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 2000: ¥4,100,000 (Direct Cost: ¥4,100,000)
Fiscal Year 1999: ¥4,900,000 (Direct Cost: ¥4,900,000)
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| Keywords | dystrophin / smooth muscle / transgenic mouse / Duchenne muscular dystrophy / blood vessel / 発現調節機構 / 遺伝子治療 / プロモーター / 血管平滑筋 / 血流異常 |
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| Research Abstract |
Duchenne muscular dystrophy (DMD) is an X-linked fatal disease caused by mutations of the gene encoding the cytoskeletal protein dystrophin. Recent studies indicate that nNOS in skeletal muscle plays a key role in the regulation of the blood flow within exercising skeletal muscle by blunting the vasoconstrictor response to alpha-vasoconstrictor response to alpha-adrenergic receptor activation. We hypothesized that dystrophin deficiency also causes the reduction of nNOS in vascular smooth muscle cells (VSMCs), leads to vascular dysfunction and exacerbates muscle pathology. We therefore generated transgenic mice expressing 14 Kb full-length human dystrophin cDNA under the transcriptional control of the smooth muscle alpha-actin promoter. These mice were then crossed with mdx mice, resulting in three independent SMTg/mdx lines which harbor the dystrophin gene only in SMCs. The expression pattern was detectable by semi-quantitative RT-PCR analysis and immunohistotochemical staining, which showed the specific expression of transgene in SMCs. We are analyzing histological characteristics of SMTg/mdx mices such as central nucleation, fiber size variability, and CK concentrations as compared to C57BL/10 control mice and mdx mice.
In addision, we have succeeded that LacZ gene was expressed in Vascular smooth muscle cell (VSMC) by RGD-modification adenovirus using muscle specific promoter for gene therapy targeting to vascular muscle of DMD.
We also developed the methods of targeted and stable gene deliverly into muscle cells by a two-step transfer.
We showed that the analysis of dystrophin expression in myotubes which were differenciated from fibloblasts is useful for genetic diagnosis of DMD
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