Spin-trapping detection of nitric oxide (NO) production from macrophage during matuation

自旋捕获检测巨噬细胞成熟过程中一氧化氮 (NO) 的产生

• 批准号: 07660387
• 负责人: KUWABARA Mikinori
• 金额: $ 1.66万
• 依托单位: HOKKAIDO UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1995
• 资助国家: 日本
• 起止时间: 1995 至 1996
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07660387/
• 关键词: Nitric oxide; Macrophage; MGD; ESR; Interferon-gamma; Lipopolysaccharide; lactoferrin; spin-trapping; 単球; 一酸化窒素; スーパーオキシド; スピントラッピング; 電子スピン共鳴法; メチルグルカミンジカルバメート

In order to investigate nitric oxide (NO) production from macrophage during maturation, peneatrial macrophage was isolated from thioglycolate-stimulated mouse and the production of NO after stimulation by interferon-gamma (INF-gamma), lipopolysaccharide (LPS) and lactoferrin (LF) was measured by the method combining electron-spin-resonance spectroscopy (ESR) and spin-trapping. The complex (MGD-Fe) of Fe^<2+> and N- (Dithiocarbonyl) -N-methylglucamine was used as spin-trapping reagent. Triplet ESR signal due to MGD-Fe-NO,which was produced by the reaction of NO and MGD-Fe, was time-dependently observed in LPS+Inf-gamma-stimulated macrophage. The NO production was also confirmed the increased-responses of NO_2^-+NO_3^-, metabolites of NO,by colorimetric method using Griess reagents. It was also shown that NO synthase detected by Western blotting was induced by these stimulation. Furthermore, LF promoted the NO production and iNOS induction in INF-gamma-stimulated macrophage, whereas LF itself did not produce any NO production in macrophage. These results was suggested that the maturation of macrophage was induced by LF which was produced from neutrophil in inflammation site as well as various cytokines which was produced from various lymphocytes.

为了研究成熟过程中巨噬细胞产生的一氧化氧化物（NO）产生，从硫甘氨酸蛋白刺激的小鼠中分离了一种巨噬细胞，并通过通过方法组合式培训来测量甲状腺酸盐刺激的小鼠（INF-GAMMA），脂多糖（INF-GAMMA），脂多糖（INF-GAMMA），脂多糖酸（LPS）和乳糖素（LF）的NO刺激后的NO产生。 Fe^<2+>和N-（二硫代）-n-甲基葡萄糖葡萄糖的复合物（MGD-FE）用作自旋捕获试剂。由NO和MGD-FE的反应产生的MGD-FE-NO引起的TripleT ESR信号在LPS+INF-GAMMA刺激的巨噬细胞中观察到时间依赖性。 NO生产也被证实了使用Griess试剂的LOLLIMETICET方法，NO_2^ - +NO_3^ - NO_2^ - +NO_3^ - 的增加。还表明，这些刺激诱导了通过蛋白质印迹检测到的合成酶。此外，LF促进了INF-GAMMA刺激的巨噬细胞中的NO生产和iNOS诱导，而LF本身并未在巨噬细胞中产生任何生产。这些结果表明，巨噬细胞的成熟是由LF诱导的，LF是由炎症部位中嗜中性粒细胞产生的，以及由各种淋巴细胞产生的各种细胞因子。

项目成果

Ohshima, H.: "Damage induced by hydroxyl radicals generated in the hydration layer of gamma-irradiated frozen aqueous solution of DNA." Jouranal of Radiation Research. 37. 199-207 (1996)

Ohshima, H.：“伽马射线照射的冷冻 DNA 水溶液的水合层中产生的羟基自由基引起的损伤。”

Kuwabara, M.: "Lipid peroxide levels and superoxide-scavenging abilities of sera obtained from hotbred (Thoroughbred) horses." Journal of Veterinary Medical Science. 58. 97-101 (1996)

Kuwabara, M.：“从纯种马获得的血清的脂质过氧化物水平和超氧化物清除能力。”

Nagahata, H., Higutchi, H., Goji, N., Noda, H.and Kuwabara, M: "Functional characteristics of enhanced Fc receptor expression of beta2 integrin-deficient bovine mononuclear phagocyte." Microbiol.Immunol.40. 389-395 (1996)

Nagahata, H.、Higutchi, H.、Goji, N.、Noda, H. 和 Kuwabara, M：“β2 整合素缺陷型牛单核吞噬细胞 Fc 受体表达增强的功能特征。”

Nagahata, H.: "Functional characteristics of enhanced Fc receptor expression of β2 integrin-deficient bovine mononuclear phagocyte." Microbiology and Immunology. 40. 389-395 (1996)

Nagahata, H.：“β2 整合素缺陷的牛单核吞噬细胞 Fc 受体表达增强的功能特征。微生物学和免疫学。” 40. 389-395 (1996)

Kuwabara, M., Inukai, N., Inanami, O., Miyake, Y., Tsunoda, N., Maki, Y.and Sato, F: "Lipid peroxide levels and superoxide-scavenging abilities of sera obtained from hotbred (Thoroughbred) horses." J.Vet.Med.Sci.58. 97-101 (1996)

Kuwabara, M.、Inukai, N.、Inanami, O.、Miyake, Y.、Tsunoda, N.、Maki, Y.和 Sato, F：“从杂交种（纯种马）获得的血清的脂质过氧化物水平和超氧化物清除能力