Sauer Heinrich, Bekhite Mohamed M, Hescheler Jürgen, Wartenberg Maria
Department of Physiology, Justus-Liebig-University Giessen, D-35392 Giessen, Germany.
Exp Cell Res. 2005 Apr 1;304(2):380-90. doi: 10.1016/j.yexcr.2004.11.026. Epub 2005 Jan 9.
The molecular mechanisms driving angiogenesis in tissues derived from embryonic stem (ES) cells are currently unknown. Herein we investigated the effects of direct current (DC) electrical field treatment on endothelial cell differentiation and angiogenesis of mouse ES cells. Treatment of ES cell-derived embryoid bodies with field strengths ranging from 250 V/m to 750 V/m, applied for 60 s, dose-dependently increased the capillary area staining positive for the endothelial-specific marker platelet endothelial cell adhesion molecule-1 (PECAM-1), indicating stimulation of endothelial cell differentiation and angiogenesis. Consequently, increased expression of hypoxia-inducible factor-1alpha (HIF-1alpha) and vascular endothelial growth factor (VEGF) within 24 h was observed. Electric field treatment raised reactive oxygen species (ROS) generation for at least 48 h, which was blunted by NADPH-oxidase inhibitors diphenylen iodonium chloride (DPI) as well as 4-(2-aminoethyl)benzenesulfonyl fluoride (AEBSF), and increased the expression of NADPH-oxidase subunits p22-phox, p47-phox, p67-phox, and gp91-phox within 24 h. Electrical field treatment resulted in activation of extracellular regulated kinase 1,2 (ERK1,2), p38, as well as c-Jun NH2-terminal kinase (JNK). Pretreatment with the JNK inhibitor SP600125 resulted in a significant decrease in capillary areas under control conditions as well as under conditions of electrical field treatment, whereas the p38 inhibitor SB203580 was without effects. By contrast, the ERK1,2 antagonist UO126 inhibited electrical field-induced angiogenesis, whereas angiogenesis under control conditions was unimpaired. The increase in capillary areas and VEGF expression as well as activation of JNK and ERK1,2 was significantly inhibited in the presence of the free radical scavenger vitamin E underscoring the role of ROS in electrical field-induced angiogenesis of ES cells.
驱动胚胎干细胞（ES细胞）来源组织中血管生成的分子机制目前尚不清楚。在此，我们研究了直流（DC）电场处理对小鼠ES细胞内皮细胞分化和血管生成的影响。用强度范围为250 V/m至750 V/m的电场处理ES细胞来源的胚状体60秒，剂量依赖性地增加了内皮特异性标志物血小板内皮细胞黏附分子-1（PECAM-1）染色阳性的毛细血管面积，表明刺激了内皮细胞分化和血管生成。因此，观察到24小时内缺氧诱导因子-1α（HIF-1α）和血管内皮生长因子（VEGF）的表达增加。电场处理使活性氧（ROS）生成至少增加48小时，这被NADPH氧化酶抑制剂二亚苯基碘鎓氯化物（DPI）以及4-（2-氨基乙基）苯磺酰氟（AEBSF）减弱，并在24小时内增加了NADPH氧化酶亚基p22-phox、p47-phox、p67-phox和gp91-phox的表达。电场处理导致细胞外调节激酶1,2（ERK1,2）、p38以及c-Jun氨基末端激酶（JNK）的激活。用JNK抑制剂SP600125预处理导致在对照条件下以及电场处理条件下毛细血管面积显著减少，而p38抑制剂SB203580没有作用。相比之下，ERK1,2拮抗剂UO126抑制电场诱导的血管生成，但对照条件下的血管生成未受影响。在存在自由基清除剂维生素E的情况下，毛细血管面积和VEGF表达的增加以及JNK和ERK1,2的激活被显著抑制，这突出了ROS在电场诱导的ES细胞血管生成中的作用。
