A novel effect of insulin on vascular smooth muscle cells through the transcriptional regulation ?the meaning of the activation of C/EBP

胰岛素通过转录调控对血管平滑肌细胞的新作用——C/EBP激活的意义

• 批准号: 14571090
• 负责人: NISHIO Yoshihiko
• 金额: $ 2.24万
• 依托单位: Shiga University of Medical Science
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2002
• 资助国家: 日本
• 起止时间: 2002 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14571090/
• 关键词: Insulin; PI3-kinase; CCAAT; enhancer binding protein; smooth muscle cells; transcription; monocyte chemoattractant protein-1; MCP-1; PI3-kinases

Phosphatidylinositol 3-kinase (PI 3-kinase is a key molecule mediating signals of insulin in vascular smooth muscle cells (VSMCs). To examine the-effect of chronic activation of PI 3-kinase on the gene expression of VSMCs, membrane-targeted p110CAAX, a catalytic subunit of PI 3-kinase, was o overexpressed m rat SMCs by adenovirus-mediated gene transfer. Similar to insulin' s effects, cells overexpressing p110CAAX exhibited 10-to 15-fold increase in monocyte chemoattractant protein-1 (MCP-1) mRNA expression as compared with the control cells. Electrophoretic mobility shift assay showed that the overexpression of p110CAAX activated neither the NF-κB binding nor the activator protein (AP-1) binding activities. To investigate the mechanisms for the induction of MCP-1 gene expression by p110CAAX, 3.6 kb of the upstream region of MCP-1 gene was cloned and the promoter activity was analyzed by luciferase reporter assay. We found that two CCAAT/enhancer binding protein (C/EBP) binding sites located between 2.6 and 3.6 kb upstream of the MCP-1 gene were responsible for the induction by p110CAAX. The overexpression of C/EBP-β and C/EBP-δ but not C/EBP-α caused 6-to 8-fold induction of MCP-1 promoter activity. Consistently, the overexpression of p110CAAX induced mRNA expression and nuclear expression of CIEBP-β and C/EBP-δ in VSMCs. Insulin at 1 to 10 nM also increased nuclear expression of C/EBP-βand C/EBP-δ in VSMCs. These results clearly indicate that the activation of PI 3-kinase induced proinflammatory gene expression through activating C/EBP-β and CIEBP-δ but not NF-κB, which may explain the proinflammatory effect of insulin in the insulin-resistant state.

磷脂酰肌醇3-激酶（PI 3-激酶）是血管平滑肌细胞中介导胰岛素信号的关键分子。为了研究PI 3激酶的慢性激活对VSMCs基因表达的影响，通过腺病毒介导的基因转移，膜靶向p110CAAX （PI 3激酶的催化亚基）在大鼠SMCs中过表达。与胰岛素的作用类似，过表达p110CAAX的细胞单核细胞趋化蛋白-1 (MCP-1) mRNA的表达比对照细胞增加了10- 15倍。电泳迁移率转移实验表明，p110CAAX过表达既没有激活NF-κB结合，也没有激活激活蛋白（AP-1）的结合活性。为了研究p110CAAX诱导MCP-1基因表达的机制，我们克隆了MCP-1基因上游3.6 kb片段，并利用荧光素酶报告基因法分析了启动子活性。我们发现两个CCAAT/增强子结合蛋白（C/EBP）结合位点位于MCP-1基因上游2.6 - 3.6 kb之间，与p110CAAX诱导有关。C/EBP-β和C/EBP-δ过表达，而C/EBP-α不表达，可诱导MCP-1启动子活性达到6 ~ 8倍。同样，p110CAAX过表达诱导VSMCs中CIEBP-β和C/EBP-δ的mRNA表达和核表达。胰岛素在1 ~ 10 nM也增加了VSMCs中C/EBP-β和C/EBP-δ的核表达。这些结果清楚地表明，PI 3激酶的激活通过激活C/EBP-β和CIEBP-δ而不是NF-κB来诱导促炎基因的表达，这可能解释了胰岛素抵抗状态下胰岛素的促炎作用。

项目成果

Shimizu S, Ugi S, Maegawa H et al.: "Protein-tyrosine phosphatase 1B as new activator for hepatic lipogenesis viasterol regulatory element-binding protein-1 gene expression"J Bilol Chem. 278. 43095-43101 (2003)

Shimizu S、Ugi S、Maekawa H 等人：“蛋白质酪氨酸磷酸酶 1B 作为肝脂肪生成维甾醇调节元件结合蛋白 1 基因表达的新激活剂”J Bilol Chem。

Sekine O, Nishio Y, Egawa K et al.: "Insulin acrivtes CCAAT/enhancer binding proteins and proinflammatory gene expression through the phosphatidylinositol 3-kinase pathway in vascular smooth moscle cells"The Journal of Biological Chemistry. 277. 36631-366

Sekine O、Nishio Y、Egawa K 等人：“血管平滑肌细胞中通过磷脂酰肌醇 3-激酶途径实现胰岛素促 CCAAT/增强子结合蛋白和促炎基因表达”《生物化学杂志》。

Sekine O, Nishio Y, Egawa K et al.: "Insulin activates CCAAT/enhancer binding proteins and proinflammatory gene expression through the phosphatidylinositol 3-kinase pathway in vascular smooth muscle cells."J Biol Chem. 277. 36631-36639 (2002)

Sekine O、Nishio Y、Egawa K 等人：“胰岛素通过血管平滑肌细胞中的磷脂酰肌醇 3-激酶途径激活 CCAAT/增强子结合蛋白和促炎基因表达。”J Biol Chem。

Nagai Y,.Nishio Y, Nakamura T et al.: "Amelioration of high fructose-induced metabolic derangements by activation of PPARalpha"Am J Physiol Endocrinol Metab. 282. E1180-E1190 (2002)

Nagai Y、Nishio Y、Nakamura T 等人：“通过激活 PPARα 改善高果糖诱导的代谢紊乱”Am J Physiol Endocrinol Metab。

Nagai Y, Nishio Y, Nakamura T et al.: "Amelioration of high fructose-induced metabolic derangements by activation of PPARalpha"American Journal Physiology, Endocrinolgy and Metabolism. 282. 1180-1190 (2002)

Nagai Y、Nishio Y、Nakamura T 等人：“通过激活 PPARα 改善高果糖诱导的代谢紊乱”美国生理学、内分泌和代谢杂志。