Signal transduction pathway in endothelial dysfunction.
Signal transduction pathway in endothelial dysfunction.
复制标题
内皮功能障碍的信号转导途径。
作者：
C. Wilasrusmee;G. Shah;S. Kittur;A. Halverson;D. Bruch;D. Kittur
背景
内皮功能障碍是脓毒症、急性呼吸窘迫综合征（ARDS）和其他感染性疾病的重要特征。以前，我们报道了一种研究内皮功能障碍的体外模型，其中内皮细胞通过在基底膜基质（Matrigel）上培养来诱导形成毛细血管网络。在这项研究中，我们确定了导致内皮细胞功能和毛细血管破坏的信号转导途径，败血症和其他感染性疾病的特点。
方法
将人主动脉内皮细胞（HAEC）在富含层粘连蛋白的基质上培养以形成毛细血管样网络。HAECs用蛋白酪氨酸磷酸酶抑制剂（原钒酸钠），磷酸肌醇-3-磷酸抑制剂（渥曼青霉素），或蛋白激酶C抑制剂（双吲哚马来酰亚胺）之前，毛细管形成或毛细管成熟后。通过计数一式三份威尔斯孔中毛细管网络的交叉点来定量毛细管形成的程度。通过方差分析确定统计学显著性。
结果
抑制蛋白酪氨酸磷酸酶或蛋白激酶C后出现内皮功能障碍。而磷酸肌醇-3-磷酸的抑制不会引起内皮功能障碍，原钒酸钠（2-20 μ M）和双吲哚马来酰亚胺（2-10 μ M）显着减少毛细血管网络。对照组、原钒酸钠处理组和双吲哚马来酰亚胺处理组中毛细血管数量的平均值+/- SD分别为251.0 +/- 7.0、65.6 +/- 9.9（p < 0.001）和181.7 +/- 0.1（p < 0.001）。原钒酸钠（20-200 μ M）和双吲哚马来酰亚胺（10-100 μ M）抑制毛细管形成。在较高浓度下，原钒酸钠（> 200 μ M）和双吲哚马来酰亚胺（>100 μ M）破坏成熟的毛细管。
结论
我们的研究结果表明，PKC和蛋白酪氨酸磷酸酶在内皮功能障碍中发挥作用，通过干扰内皮细胞内的磷酸化信号。这些机制可能在败血症和其他感染性疾病的内皮功能障碍中起重要作用。
BACKGROUND
Endothelial dysfunction is an important feature of sepsis, acute respiratory distress syndrome (ARDS), and other infectious conditions. Previously, we reported an in vitro model to study endothelial dysfunction, in which endothelial cells are induced to form capillary tube networks by culturing on a basement membrane matrix (Matrigel). In this study, we defined the signal transduction pathways that lead to endothelial cell function and capillary disruption characteristic of sepsis and other infectious conditions.
METHODS
Human aortic endothelial cells (HAEC) were cultured on a laminin-rich matrix to form capillary-like networks. The HAECs were treated with a protein tyrosine phosphatase inhibitor (sodium orthovanadate), a phosphoinositon-3-phosphate inhibitor (wortmannin), or a protein kinase C inhibitor (bisindolylmaleimide) before capillary tubes had formed or after the capillary tubes had matured. The degree of capillary tube formation was quantified by counting the intersection of capillary networks in triplicate wells. Statistical significance was determined by analysis of variance.
RESULTS
Endothelial dysfunction occurred after inhibition of protein tyrosine phosphatase or protein kinase C. Whereas inhibition of phosphoinositon-3-phosphate did not cause endothelial dysfunction, sodium orthovanadate (2-20 microM) and bisindolylmaleimide (2-10 microM) significantly reduced capillary networks. The mean +/- SD of the number of capillary tubes in the control, sodium orthovanadate-treated, and bisindolylmaleimide-treated groups were 251.0 +/- 7.0, 65.6 +/- 9.9 (p < 0.001), and 181.7 +/- 0.1 (p < 0.001), respectively. Sodium orthovanadate (20-200 microM) and bisindolylmaleimide (10-100 microM) inhibited capillary tube formation. At higher concentrations, sodium orthovanadate (> 200 microM) and bisindolylmaleimide (>100 microM) disrupted mature capillary tubes.
CONCLUSIONS
Our results suggest that PKC and protein tyrosine phosphatase play a role in endothelial dysfunction by interfering with the phosphorylation signals within endothelial cells. These mechanisms may be important in the endothelial dysfunction in sepsis and other infectious conditions.