Department of Intensive Care Medicine, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland.
Liver Int. 2009 Nov;29(10):1582-92. doi: 10.1111/j.1478-3231.2009.02109.x. Epub 2009 Sep 9.
BACKGROUND/AIMS: Genes encoding for some of the mitochondrial proteins are under the control of the transcriptional factor hypoxia inducible factor-1 alpha (HIF-1 alpha), which can accumulate under normoxic conditions in inflammatory states. The aim of this study was to evaluate the effects of cobalt chloride (CoCl(2), a hypoxia mimicking agent), tumour necrosis factor-alpha (TNF-alpha) and toll-like receptor (TLR) -2, -3 and -4 agonists on HIF-1 alpha accumulation, and further on HIF-1 alpha-mediated modulation of mitochondrial respiration in cultured human hepatocytes.
The human hepatoma cell line HepG2 was used in this study. Cells were treated with CoCl(2), TNF-alpha and TLR-2, -3 and -4 agonists. HIF-1 alpha was determined by Western blotting and mitochondrial respiration in stimulated cells by high-resolution respirometry.
CoCl(2), TNF-alpha and TLR agonists induced the expression of HIF-1 alpha in a time-dependent fashion. TNF-alpha and CoCl(2), but not TLR agonists, induced a reduction in complex I-, II- and IV-dependent mitochondrial oxygen consumption. TNF-alpha-associated reduction of cellular oxygen consumption was abolished through inhibition of HIF-1 alpha activity by chetomin (CTM). Pretreatment with cyclosporine A prevented CoCl(2)-induced reduction of complex I- and II-dependent mitochondrial oxygen consumption and TNF-alpha-induced reduction of complex-I-dependent respiration, implicating the involvement of the mitochondrial permeability transition pore openings. TNF-alpha and TLR-2, -3 and -4 agonists induced the expression of vascular endothelial growth factor, which was partially abolished by the blockage of HIF-1 alpha with CTM.
The data suggest that HIF-1 alpha modulates mitochondrial respiration during CoCl(2) and TNF-alpha stimulation, whereas it has no effect when induced with TLR-2, -3 and -4 agonists.
背景/目的：一些线粒体蛋白的编码基因受转录因子缺氧诱导因子-1α（HIF-1α）的控制，在炎症状态下的常氧条件下，HIF-1α可以积累。本研究旨在评估氯化钴（CoCl2，一种模拟缺氧的试剂）、肿瘤坏死因子-α（TNF-α）和 Toll 样受体（TLR）-2、-3 和 -4 激动剂对 HIF-1α积累的影响，以及进一步对培养的人肝细胞中线粒体呼吸的 HIF-1α介导的调节作用。
本研究使用人肝癌细胞系 HepG2。用 CoCl2、TNF-α 和 TLR-2、-3 和 -4 激动剂处理细胞。通过 Western 印迹法测定 HIF-1α，并用高分辨率呼吸仪测定刺激细胞中的线粒体呼吸。
CoCl2、TNF-α 和 TLR 激动剂以时间依赖的方式诱导 HIF-1α的表达。TNF-α和 CoCl2，但不是 TLR 激动剂，诱导依赖于复合物 I、II 和 IV 的线粒体耗氧量减少。通过 chetomin（CTM）抑制 HIF-1α活性，TNF-α 相关的细胞耗氧量减少被消除。环孢菌素 A 的预处理可防止 CoCl2 诱导的复合物 I 和 II 依赖性线粒体耗氧量减少和 TNF-α 诱导的依赖于复合物 I 的呼吸减少，这表明涉及线粒体通透性转换孔的开放。TNF-α和 TLR-2、-3 和 -4 激动剂诱导血管内皮生长因子的表达，用 CTM 阻断 HIF-1α部分消除了这种表达。
数据表明，在 CoCl2 和 TNF-α刺激期间，HIF-1α调节线粒体呼吸，而在用 TLR-2、-3 和 -4 激动剂诱导时则没有影响。
