| Project/Area Number |
15591404
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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 |
Digestive surgery
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| Research Institution | Osaka University |
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Principal Investigator |
UMESHITA Koji Osaka University, Hospital, Assistant Professor, 医学部附属病院, 助手 (60252649)
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| Co-Investigator(Kenkyū-buntansha) |
KANEDA Yasufumi Osaka University, Graduate School of Medicine, Professor, 医学系研究科, 教授 (10177537)
NAGANO Hiroaki Osaka University, Graduate School of Medicine, Associate Professor, 医学系研究科, 講師 (10294050)
左近 賢人 大阪大学, 医学系研究科, 助教授 (40170659)
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| Project Period (FY) |
2003 – 2004
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| Project Status |
Completed (Fiscal Year 2004)
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| Budget Amount *help |
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2004: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 2003: ¥2,100,000 (Direct Cost: ¥2,100,000)
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| Keywords | Liver / ischemia-reperfusion / transplantation / gene therapy / RNA / electroporation / HVJ Envelope Vector / 遺伝子導入 / 肝移植 / ex vivo / 虚血・再潅流傷害 / RNA導入 / HVJエンベロープベクター / 蛋白導入 / マウス |
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| Research Abstract |
In the first year of the study period, we succeeded in stable synthesis and amplification of mRNA using mCAP RNA Capping Kit. Next, we moved to in vitro mRNA transfer study using HVJ Envelope VECTOR KIT and got 500- to 1000-fold increase in luciferase assay in BHK21 cells compared with control. When HVJ envelope vector containing luciferase mRNA was injected to the liver via portal vein in C57BL/6 mice in vivo, 5- to 10-fold increase was obtained. In the second year, we injected mRNA to the liver via portal vein in Wistar rats at 4C ex vivo using the same method. However, only 2- to 3-fold increase was observed in luciferase assay. We, therefore, tried ultrasound-mediated gene transfer with echo contrast microbubble (Optison), which has recently been shown to be highly efficient, in stead of HVJ envelope vector. In the preliminary experiment using luciferase reporter gene, luciferase activity of liver graft 24 hours after transfection and syngeneic transplantation was significantly increased about 17-fold as compared with plasmid infusion alone. When vascular clamping was performed together, additional 10-fold increase was obtained. In the next step, mRNA transfer into liver graft using ultrasound-mediated method would be performed.
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