| Project/Area Number |
16590249
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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 |
Pathological medical chemistry
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| Research Institution | Kumamoto University |
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Principal Investigator |
AKUTA Teruo Kumamoto University, Graduate School of Medical Sciences, Assistant Professor, 大学院・医学薬学研究部, 助手 (00346975)
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| Co-Investigator(Kenkyū-buntansha) |
AKAIKE Takaaki Kumamoto University, Graduate School of Medical Sciences, Professor, 大学院・医学薬学研究部, 教授 (20231798)
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| Project Period (FY) |
2004 – 2005
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| Project Status |
Completed (Fiscal Year 2005)
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| Budget Amount *help |
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2005: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 2004: ¥2,000,000 (Direct Cost: ¥2,000,000)
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| Keywords | NO / 8-nitroguanosine / cGMP / 8-nitroc-GMP / anti-8-nitro-cGMP antibody / protein kinase G / cell signaling / redox signaling / 抗8-ニトログアノシン抗体 / 細胞保護作用 / ヘムオキシゲナーゼ-1 / RNAウイルス / 遺伝子変異 |
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| Research Abstract |
We examined biological and signal function of a novel nucleic acids modification by NO, especially, 8-nitroguanosine (8-NO_2Guo) and 8-NO_2Guo-related compounds, e.g. 8-nitro-cyclic GMP (8-NO_2-cGMP). Our data on these compounds are summarized as follows.
1) 8-NO_2Guo formation was mainly localized in cytosol, near the endoplasmic reticulum, in HepG2 cells treated with NO, as identified by confocal microscopy and immunoelectron microscopy analysis using anti 8-NO_2Guo monoclonal antibody. We demonstrated that 8-NO_2Guo protected from apoptosis HepG2 cell through upregulation of heme oxygenase-1 and endothelial NOS (eNOS), and inhibition of bcl-XL degradation.
2) 8-NO_2Guo has mutagenic potential in DNA and RNA. Authentic 8-NO_2Guo added exogenously to cultured CV-1 cells infected with a recombinant Sendai virus containing a green fluorescent protein gene increased the viral mutation frequency. This mutation spectrum was dominantly observed with a C to U transition. 8-NO_2Guo added to AS5
… More
2 cells also increased the genomic mutation frequency, with a predominant mutation, G to T transition. We supposed two different mechanisms for this mutagenesis : direct modification of nucleic acid and indirect augmentation of oxidative stress via superoxide generation by 8-NO_2Guo.
3) We established the synthesis method of 8-NO_2-cGMP and prepared large amount of this compound. Treatment of human uterine smooth muscle cells with 8-NO_2-cGMP increased in cGMP-dependent protein kinase-associated phosphorylation of vasodilator-stimulated phosphoprotein on Ser^<239>. Strong vasorelaxing potential for rat carotid artery was observed with 8-NO_2-cGMP. These results suggested that 8-NO_2-cGMP is a new cellular redox signaling molecule.
4) We developed the anti-8-NO_2-cGMP monoclonal and polyclonal antibodies. The immunostaining with anti-8-NO_2-cGMP monoclonal antibody showed high immuoreactivity in cytosol of mouse macrophage derived RAW264 cells stimulated with IFN-γ and lipopolysaccharide.
Taken together, 8-NO_2Guo and 8-NO_2-cGMP have unique biochemical and pharmacological properties such as cellular redox signaling and mutagenic potential. Less
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