Leibniz-Institut für Umweltmedizinische Forschung, Düsseldorf, Germany.
Horm Metab Res. 2011 Apr;43(4):268-74. doi: 10.1055/s-0031-1271692. Epub 2011 Feb 17.
The phosphoinositide 3'-kinase (PI3 K)/Akt pathway controls the activity of a number of proteins important in the regulation of apoptosis and cell proliferation. FoxO (forkhead box, class O) transcription factors, substrates of the Ser/Thr kinase Akt, control the expression of several target genes that are crucial to the defense against oxidative stress, the regulation of cell cycle, and apoptosis in mammalian cells. Here, expression of ceruloplasmin (CP), the major copper-containing protein in blood released by the liver, was investigated. We observed a significant downregulation of CP mRNA levels after insulin treatment in H4IIE rat hepatoma cells. The PI3K inhibitor wortmannin counteracted this insulin effect on CP mRNA levels, indicating that the PI3K/Akt cascade is involved in the regulation of CP expression. Stimulation of FoxO1 was induced in H4IIE rat hepatoma cells expressing a conditionally active FoxO1 construct, resulting in significant upregulation of CP mRNA levels. This upregulation was prevented in the presence of insulin. In parallel, mRNAs of established FoxO target genes were analyzed: like CP mRNA, selenoprotein P and glucose 6-phosphatase mRNAs were upregulated by FoxO1, which was prevented by insulin. The same effects of insulin on CP mRNA levels were detected in primary rat hepatocytes. Furthermore, CP release into cell culture media was analyzed with primary hepatocytes and found to be attenuated by insulin. In line with its insulin-mimetic effects on cultured cells, Cu (2+) imitated the effect of insulin on CP expression and caused a downregulation of CP mRNA levels in rat hepatoma cells.
磷酸肌醇 3-激酶（PI3K）/Akt 途径控制着许多在调节细胞凋亡和增殖中起重要作用的蛋白质的活性。FoxO（叉头框，O 类）转录因子是 Akt 丝氨酸/苏氨酸激酶的底物，控制着几种靶基因的表达，这些基因对哺乳动物细胞的氧化应激防御、细胞周期调节和细胞凋亡至关重要。在这里，我们研究了铜蓝蛋白（CP）的表达，CP 是肝脏释放的血液中主要的含铜蛋白。我们观察到胰岛素处理 H4IIE 大鼠肝癌细胞后 CP mRNA 水平显著下调。PI3K 抑制剂wortmannin 拮抗了胰岛素对 CP mRNA 水平的这种作用，表明 PI3K/Akt 级联参与 CP 表达的调节。在表达条件性激活 FoxO1 构建体的 H4IIE 大鼠肝癌细胞中诱导 FoxO1 刺激，导致 CP mRNA 水平显著上调。在胰岛素存在的情况下，这种上调被阻止。平行地，分析了已建立的 FoxO 靶基因的 mRNAs：像 CP mRNA 一样，硒蛋白 P 和葡萄糖 6-磷酸酶 mRNAs 被 FoxO1 上调，而胰岛素则阻止了这种上调。在原代大鼠肝细胞中也检测到了胰岛素对 CP mRNA 水平的相同影响。此外，用原代肝细胞分析了 CP 向细胞培养物培养基中的释放情况，发现胰岛素减弱了 CP 的释放。与它在培养细胞上的胰岛素模拟作用一致，Cu（2+）模拟了胰岛素对 CP 表达的作用，并导致大鼠肝癌细胞中 CP mRNA 水平下调。
