Development of a simple screening system for the discovery of a new drug for diabetes

开发用于发现糖尿病新药的简单筛选系统

• 批准号: 10557019
• 负责人: EBINA Yousuke
• 金额: $ 8.38万
• 依托单位: The University of Tokushima
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B).
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-10557019/
• 关键词: Type 2 diabetes; Serumsubstance caused for insulin resistance; インスリン抵抗性; NIDDM; トランスジェニックラット; GLUT4myc; インスリン非依存型糖尿病

NIDDM is primarily caused by the insulin resistance, and increase insulin secretion from pancreatic β-cells and sub-sequently deficiency of insulin. Therefore it is a key to find a substance for insulin resistance in the blood of the patient. There are several substrates in the plasma for insulin resistance, TGF-β, leptin and resistine. However, they are uncertain for insulin resistance of the onset of NIDDM.Translocation of the type 4 glucose transporter (GLUT4) to the cell surface from an intracellular pool is the major mechanism of insulin-stimulated glucose uptake in insulin-target cells which is one of the most important physiological effect of insulin. We developed a highly sensitive and quantitative method to detect GLUT4 immunologically on the surface of intact cells, using c-myc epitope-tagged GLUT4 (GLUT4myc). We established stable clones to express GLUT4myc in 3T3L1 adipocytes and established the transgenic rat expressing GLUT4myc in rat adipocytes. We have developed a simple and new method to screen the compounds for triggering the GLUT4myc translocation in 96 wells using luminescence method. This method is useful for the first screening for finding the substances for insulin resistance. Recently, we found a part of known molecule may be the substance for insulin resistance.

NIDDM主要是由胰岛素抵抗引起的，并增加胰腺β细胞的胰岛素分泌，从而导致胰岛素缺乏。因此寻找患者血液中胰岛素抵抗的物质是关键。血浆中存在多种胰岛素抵抗底物、TGF-β、瘦素和抵抗素。然而，他们对于NIDDM发病的胰岛素抵抗尚不确定。4型葡萄糖转运蛋白(GLUT4)从细胞内池易位至细胞表面是胰岛素刺激靶细胞中葡萄糖摄取的主要机制，这是胰岛素最重要的生理效应之一。我们开发了一种高度灵敏的定量方法，使用 c-myc 表位标记的 GLUT4 (GLUT4myc) 免疫学检测完整细胞表面的 GLUT4。我们建立了在3T3L1脂肪细胞中表达GLUT4myc的稳定克隆，并建立了在大鼠脂肪细胞中表达GLUT4myc的转基因大鼠。我们开发了一种简单而新的方法，利用发光法在 96 孔中筛选触发 GLUT4myc 易位的化合物。该方法对于首次筛选寻找胰岛素抵抗物质是有用的。最近，我们发现部分已知分子可能是胰岛素抵抗的物质。

项目成果

T.Imanaka, Y.Ebina et. al.: "Reconstitution of Insulin Signaling Pathways in Rat 3Y1 Cells Lacking Insulin Receptor and Insulin Receptor Substrate-1"J.Biol.Chem.. 273. 25347-25355 (1998)

T.Imanaka，Y.Ebina 等。

Oleg V.Chaika Yousuke Ebina,et al.: "Mutation of Tyrosine 960 within the Insulin Receptor Juxtamembrane Domain Impairs Glucose Transport but Dose Not Inhibit Ligand-mediated Phosphorylation of Insulin Recepte-2 tor Substra in 3T3-L1 Adipocytes"J.Biol.Chem

Oleg V.Chaika Yousuke Ebina 等人：“胰岛素受体近膜结构域内酪氨酸 960 的突变会损害葡萄糖转运，但剂量不会抑制 3T3-L1 脂肪细胞中胰岛素受体 2 的配体介导的磷酸化”J.Biol。

L.Wang, Y.Ebina et. al.: "Gi-mediated translocation of GLUT4 is independent of p85/p110α and p110γ phosphoinositide 3-kinases but might involve the activation of Akt kinase"Biochem.J.. 345. 543-555 (2000)

L.Wang、Y.Ebina 等人：“Gi 介导的 GLUT4 易位独立于 p85/p110α 和 p110γ 磷酸肌醇 3-激酶，但可能涉及 Akt 激酶的激活”Biochem.J. 345. 543-555 (2000)

Hiroyuki Sano,Yousuke Ebina,et al.: "Insulin enhances macrophage scavenger receptor-mediated endocytic uptake of advanced glycation and products" J.Biol.Chem.273. 8630-8637 (1998)

Hiroyuki Sano、Yousuke Ebina 等人：“胰岛素增强巨噬细胞清道夫受体介导的晚期糖基化和产物的内吞摄取”J.Biol.Chem.273。

Lihong Wang,Yousuke Ebina,et al.: "Transient Effect of Platelet-derived Growth Factor(PDGF) on GLUT4 Translocation in 3T3-L1 Adipocytes"J.Biol.Chem.. 274. 19246-19253 (1999)

Lihong Wang,Yousuke Ebina,et al.:“血小板衍生生长因子（PDGF）对 3T3-L1 脂肪细胞中 GLUT4 易位的瞬时影响”J.Biol.Chem.. 274. 19246-19253 (1999)