Studies on the suppressive factor from parasite of NO synthase and cytokine gene expression in macrophages

寄生虫NO合酶抑制因子及巨噬细胞细胞因子基因表达的研究

• 批准号: 11670241
• 负责人: FUKUMOTO Soji
• 金额: $ 2.3万
• 依托单位: TOTTORI UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11670241/
• 关键词: Spriometra erinaceieuropaei; macrophage; RAW 264.7; TNF-α; nitric oxide synthase; chemokine; gene expression; NF-κB; IL-1; Northern blot

We found that excretory/secretory (ES) products of Spirometra erinaceieuropaei suppress the gene expression of iNOS, TNF-α, chemokines in mouse peritoneal macrophages stimulated with LPS.We examined the suppesive mechanism of gene expression in the present investigation.(1) The TNF-α gene expression was suppressed by ES products downstream of TLR4 based on the comparison of the effect of ES products in C3H/HeN miceand C3H/HeJ mice, in which point mutation was observed in TLR4.(2) These suppressive effects were not related with prostaglandin E2, IL-10 and secretory leukocyte protease inhibitor (SLPI) , which are produced in macrophages and suppress the macrophage functions.(3) The stability of TNF-αmRNA was not reduced by the treatment of ES products.(4) The translocation of NF-κB to nucleus and NF-κB binding activity to κB element was not reduced by the treatment of ES products.(5) In J774.1 and RAW 264.7 cell line macrophages, nitric oxide production and gene expression of IP-10 and TNF-α were suppressed. Then, we found that these cell lines are useful in the studies of suppressive mechanism.(6) ES inhibited the phospho-p38 MAP kinase and phospho-ERK as well as MAPK inhibitors, SB 203580 and PD 98059, respectively. We supposed that this is one of the suppressive mechanism of gene expression.(7) The suppressive effect of ES products almost diminished by the treatment with trypsin 15 μg/ml or protease K0.1U/ml. Judging from these data, the supressive factor is supposed to be a protein.

我们发现，Spirometra erinaceieuropaei 的排泄/分泌（ES）产物抑制 LPS 刺激的小鼠腹腔巨噬细胞中 iNOS、TNF-α、趋化因子的基因表达。我们在本研究中检查了基因表达的抑制机制。（1）基于 ES 产品在 C3H/HeN 小鼠中的效果比较，TLR4 下游的 ES 产品抑制了 TNF-α 基因表达C3H/HeJ小鼠，在TLR4中观察到点突变。(2)这些抑制作用与巨噬细胞产生并抑制巨噬细胞功能的前列腺素E2、IL-10和分泌性白细胞蛋白酶抑制剂(SLPI)无关。 (3)ES产物的处理并未降低TNF-αmRNA的稳定性。(4)NF-κB向细胞核的易位以及NF-κB与细胞核的结合活性。 ES产品的处理并没有减少κB元件。(5)在J774.1和RAW 264.7细胞系巨噬细胞中，一氧化氮的产生以及IP-10和TNF-α的基因表达受到抑制。 (6)ES抑制磷酸p38 MAP激酶和磷酸ERK以及MAPK抑制剂SB分别为203580和PD 98059，我们推测这是基因表达的抑制机制之一。(7)判断ES产物的抑制作用在胰蛋白酶15μg/ml或蛋白酶K0.1U/ml处理后几乎减弱。从这些数据来看，抑制因子应该是一种蛋白质。

项目成果

K.Miura, S.Fukumoto, P.Dirgahayu, K.Hirai: "Excretory/secretory products from plerocercoids of Spirometra erinaceieuropaei suppress gene expressions and production of tumor necrosis factor-αin murine macrophages stimulated with lipopolysaccharide or lipot

K.Miura、S.Fukumoto、P.Dirgahayu、K.Hirai：“来自 erinaceieuropaei Spirometra erinaceieuropaei 的 plerocercoids 的排泄/分泌产物抑制用脂多糖或 lipot 刺激的小鼠巨噬细胞中的基因表达和肿瘤坏死因子 -α 的产生

K.Miura(三浦憲豊): "Excretory/secretory products from plerocercoids of Spirometra erinaceieuropaei suppress gene expressions and production of tumor necrosis factor-α in murine macrophages stimulated with lipopolysaccharide or lipoteichoic acid. "Internationa

K.Miura（Koritoyo Miura）：“来自 Spirometra erinaceieuropaei 的 plerocercoids 的排泄/分泌产物抑制脂多糖或脂磷壁酸刺激的小鼠巨噬细胞中的基因表达和肿瘤坏死因子-α的产生。”

Haoran Wang: "Diffential expression and regulation of chemokines JE,KC,and IP-10 gene in primary cultured murine hepatocytes"Journal of Cellular Physiology. 181(2). 361-370 (1999)

王浩然：“趋化因子JE、KC和IP-10基因在原代培养的小鼠肝细胞中的差异表达和调控”细胞生理学杂志。