Armoni Michal, Harel Chava, Bar-Yoseph Fabiana, Milo Simcha, Karnieli Eddy
Institute of Endocrinology, Diabetes and Metabolism, Rambam Medical Center and B. Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa 31096, Israel.
J Biol Chem. 2005 Oct 14;280(41):34786-95. doi: 10.1074/jbc.M502740200. Epub 2005 Aug 10.
Hyperlipidemia (HL) impairs cardiac glucose homeostasis, but the molecular mechanisms involved are yet unclear. We examined HL-regulated GLUT4 and peroxisome proliferator-activated receptor (PPAR) gamma gene expression in human cardiac muscle. Compared with control patients, GLUT4 protein levels were 30% lower in human cardiac muscle biopsies from patients with HL and/or type 2 diabetes mellitus, whereas GLUT4 mRNA levels were unchanged. PPARgamma mRNA levels were 30-50% lower in patients with HL and/or diabetes mellitus type 2 than in controls. Reporter studies in H9C2 cardiomyotubes showed that HL in vitro, induced by high levels of arachidonic (AA) stearic, linoleic, and oleic acids (24 h, 200 mum) repressed transcription from the GLUT4 promoter; AA also repressed transcription from the PPARgamma1 and PPARgamma2 promoters. Co-expression of PPARgamma2 repressed GLUT4 promoter activity, and the addition of AA further enhanced this effect. 5'-Deletion analysis revealed three GLUT4 promoter regions that accounted for AA-mediated effects: two repression-mediating sequences at -443/-423 bp and -222/-197 bp, the deletion of either or both of which led to a partial derepression of promoter activity, and a third derepression-mediating sequence at -612/-587 bp that was required for sustaining this derepression effect. Electromobility shift assay further shows that AA enhanced binding to two of the three regions of cardiac nuclear protein(s), the nature of which is still unknown. We propose that HL, exhibited as a high free fatty acid level, modulates GLUT4 gene expression in cardiac muscle via a complex mechanism that includes: (a) binding of AA mediator proteins to three newly identified response elements on the GLUT4 promoter gene and (b) repression of GLUT4 and the PPARgamma genes by AA.
高脂血症（HL）会损害心脏葡萄糖稳态，但其涉及的分子机制尚不清楚。我们研究了HL对人心脏肌肉中葡萄糖转运蛋白4（GLUT4）和过氧化物酶体增殖物激活受体（PPAR）γ基因表达的调控作用。与对照患者相比，HL和/或2型糖尿病患者的人心脏肌肉活检组织中GLUT4蛋白水平降低了30%，而GLUT4 mRNA水平未发生变化。HL和/或2型糖尿病患者的PPARγ mRNA水平比对照组低30 - 50%。在H9C2心肌管中的报告基因研究表明，体外HL由高水平的花生四烯酸（AA）、硬脂酸、亚油酸和油酸诱导（24小时，200μM）可抑制GLUT4启动子的转录；AA还可抑制PPARγ1和PPARγ2启动子的转录。PPARγ2的共表达可抑制GLUT4启动子活性，添加AA可进一步增强这种作用。5'-缺失分析揭示了三个GLUT4启动子区域介导了AA的作用：位于-443/-423 bp和-222/-197 bp的两个抑制介导序列，其中任何一个或两个的缺失都会导致启动子活性部分去抑制，以及位于-612/-587 bp的第三个去抑制介导序列，该序列是维持这种去抑制作用所必需的。电泳迁移率变动分析进一步表明，AA增强了与心脏核蛋白三个区域中两个区域的结合，其性质仍不清楚。我们提出，以高游离脂肪酸水平表现的HL通过一种复杂机制调节心脏肌肉中GLUT4基因的表达，该机制包括：（a）AA介导蛋白与GLUT4启动子基因上三个新鉴定的反应元件结合；（b）AA对GLUT4和PPARγ基因的抑制作用。
