| Project/Area Number |
10670050
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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 |
General physiology
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| Research Institution | Fujita Health University |
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Principal Investigator |
OTA Akira Fujita Health Unicweairy School of Medicine Professor, 医学部, 教授 (10247637)
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| Co-Investigator(Kenkyū-buntansha) |
MORI Keiji Fujita Health Unicweairy School of Medicine Assistant Professor, 医学部, 助手 (40239596)
NAKASHIMA Akira Fujita Health Unicweairy School of Medicine Lecturer, 医学部, 講師 (20180276)
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| Project Period (FY) |
1998 – 1999
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| Project Status |
Completed (Fiscal Year 1999)
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| Budget Amount *help |
¥3,300,000 (Direct Cost: ¥3,300,000)
Fiscal Year 1999: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1998: ¥2,500,000 (Direct Cost: ¥2,500,000)
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| Keywords | NIE-115 cell line / 6R-tetrahydrobjopterin / lipopolysaccharide / GTP cyclohydrolase "I" / CD14 / CD18 / Toll-like receptor / tumor necrotizing factorα / リポポリサッカライド受容体 / N1E-115細胞 / IκB / IκBキナーゼ / 一酸化窒素 / 一酸化窒素合成酵素 |
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| Research Abstract |
6R-Tetrahydrobiopterin (BH4) is a naturally occurring essential cofactor for aromatic amino acid hydroxylases and nitric oxide (NO) synthase (NOS). Lipopolysaccharide(LPS), a component of the outer membrane of gram-negative bacilli, has been shown to enhance the BH4 content of the murine neuroblastoma cell line N1E-115 as well as the release of BH4 into culture medium. Such alterations in BH4 production were at first caused by the enhancement of the expression level of RNA encoding GTP cyclohydrolase "I"(GTPCH), a rate-limiting enzyme in the BH4 de novo biosynthetic pathway. Moreover, the expression levels of mRNAs encoding 6-pyruvoyltetrahydropterin synthase and sepiapterin reductase which place at the 2nd and the final step in BH4 biosynthesic pathway, respectively. However, we found no evidence in N1E-115 cells showing the expression of mRNA encoding CD14. CD14 is an membrane-anchoring protein and it is already recognized as the LPS receptor on macrophages. Therefore we searched the candidates for LPS receptor except for CD14 in N1E-115 cells. In a series of experiments, we found the expressions of mRNA encoding CD18, another subtype of LPS receptor on macrophages, and type 4 Toll-like receptor (T1r-4) in this cell line by way of reverse transcriptase-polymerase chain reaction. This should be the first evidence that neuron-derived cell line expresses T"I"r-4..
Tumor necrotizing factor α(TNF-α) was also capable of enhancing BH4 production in N1E-115 cells. It is currently under investigation in my laboratory whether the increase in BH4 production induced by TNF-α was caused by the activation of the pathway common to the LPS stimulation.
In spite of the increase in BH4 production by LPS stimulation. we could detect neither the increase in NO production nor the increase in the expression levels of mRNAs encoding neuronal and inducible NOS in N1E-115 cells. NOS enzymes in these cell lines were speculated to be saturated by BH4.
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