Ziegelstein Roy C, He Chaoxia, Hu Qinghua
Johns Hopkins Bayview Medical Center, Johns Hopkins University School of Medicine, Baltimore, MD 21224-2780, USA.
Biochem Biophys Res Commun. 2004 Sep 10;322(1):68-73. doi: 10.1016/j.bbrc.2004.07.080.
Increased endothelial ICAM-1 expression is found in normal aging and in atherosclerosis and is related to the chronic effects of oxidative stress. We examined the Ca(2+)-dependence of ICAM-1 mRNA expression in human aortic endothelial cells (HAEC) exposed to hypoxia/reoxygenation (H/R) as a model of oxidative stress. HAEC were exposed to glucose-free hypoxia (95% N(2)/5% CO(2)) for 60 min and were then reoxygenated (21% O(2)/5% CO(2)) and observed for up to 6h. Reactive oxygen species (ROS) generation was measured by dichlorofluorescein fluorescence and ICAM-1 mRNA was assessed by Northern blot. Upon reoxygenation after hypoxia, ROS production occurred in HAEC and was inhibited by diphenyleneiodonium and by polyethylene glycol-catalase, suggesting the involvement of NADPH oxidase-derived hydrogen peroxide. Hypoxia alone did not increase either ROS production or ICAM-1 mRNA levels, but a 2.5-fold increase in ICAM-1 mRNA was noted by 30 min of reoxygenation. This was not observed in Ca(2+)-free buffer or in cells treated with diphenyleneiodonium. Thus, H/R upregulates ICAM-1 mRNA in HAEC by a Ca(2+)- and ROS-dependent mechanism. Characterizing the signaling pathways involved in H/R-induced adhesion molecule expression may result in a better understanding of the vascular biology of normal aging and the pathobiology of atherosclerosis.
在内皮细胞中，细胞间黏附分子-1（ICAM-1）表达增加见于正常衰老和动脉粥样硬化过程，且与氧化应激的慢性效应相关。我们将缺氧/复氧（H/R）作为氧化应激模型，检测了人主动脉内皮细胞（HAEC）中ICAM-1 mRNA表达的钙依赖性。将HAEC置于无糖缺氧环境（95% N₂/5% CO₂）中60分钟，然后进行复氧（21% O₂/5% CO₂），并观察长达6小时。通过二氯荧光素荧光法测定活性氧（ROS）生成量，通过Northern印迹法评估ICAM-1 mRNA。缺氧后复氧时，HAEC中出现ROS生成，且二苯基碘鎓和聚乙二醇-过氧化氢酶可抑制其生成，提示NADPH氧化酶衍生的过氧化氢参与其中。单独缺氧既不增加ROS生成量也不增加ICAM-1 mRNA水平，但复氧30分钟时ICAM-1 mRNA增加了2.5倍。在无钙缓冲液或用二苯基碘鎓处理的细胞中未观察到此现象。因此，H/R通过钙和ROS依赖性机制上调HAEC中ICAM-1 mRNA的表达。明确H/R诱导黏附分子表达所涉及的信号通路，可能有助于更好地理解正常衰老的血管生物学和动脉粥样硬化的病理生物学。
