STUDY OF ADHESION MOLECULES, CYTOSKELETON AND SIGNAL TRANSDUCTION PATHWAY ON INJURY OF ENDOTHELIAL CELLS FOR ISOLATED PRESERVED LUNGS

离体保存肺内皮细胞损伤的粘附分子、细胞骨架及信号转导通路研究

基本信息

批准号：
10470268
负责人：
TANITA Tatsuo
金额：
$ 8.51万
依托单位：
TOHOKU UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B).
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 2000
项目状态：
已结题

项目摘要

We showed that neutrophils-induced lung injury was induced by reactive oxygen species generated via xanthine oxidase (XO) following adhesion of neutrophils on the endothelial cells. Usually, XO does not exist in the neutrophils but endothelial cells. Therefore we speculate if this kind of lung injury is induced by activation of endothelial cells themselves. In the experiments of isolated perfused rat lungs, whether intracellular signal transduction systems were involved in the injury of the pulmonary vascular endothelial cells caused by activated neutrophils, we investigated using a protein kinase C antagonist. We showed that PKC antagonist ameliorated the pulmonary vascular injury caused by activated neutrophils in a dose dependent manner. Moreover, we showed that the pulmonary vascular permeability was increased by a PKC agonist without neutrophils. On the other hand, in cell culture models, it was impossible to measure protein permeability for cell culture sheets, we measured trans- … More endothelial electrical resistance (TER) across the cultured pulmonary vascular endothelial cell sheets. We continuously measured TER of the cultured cell sheets from rat pulmonary vascular endothelial cells when they were confluent and the tested the several kinds of agonists and antagonists to the cultured cell sheets. The TERs of the cultured cell sheets were decreased by PKC agonists by dose dependent manner, meanwhile DMSO, a vehicle of PKC agonist, failed. These findings supported the results from the experiment of isolated rat lungs. We also stained actin fibers in the pulmonary vascular endothelial cells. Actin fibers existed at the peripheral area of the cytosol without any stimulations, however, they were depolymerized and moved around nucleus by PKC agonist. These findings suggest that the pulmonary vascular endothelial cells were changed their morphology by depolymerizing actin fibers by PKC when they were activated by some stimulations, and finally pulmonary microvascular permeability was increased. Less

我们发现中性粒细胞诱导的肺损伤是由中性粒细胞粘附在内皮细胞上后通过黄嘌呤氧化酶（XO）产生的活性氧引起的，因此我们推测是否存在这种情况。肺损伤是由内皮细胞本身的激活引起的。在离体灌注大鼠肺的实验中，细胞内信号转导系统是否参与了肺损伤。针对活化的中性粒细胞引起的血管内皮细胞，我们使用蛋白激酶 C 拮抗剂进行了研究，结果表明 PKC 拮抗剂以剂量依赖性方式改善了活化的中性粒细胞引起的肺血管损伤。此外，我们还发现肺血管通透性增加。另一方面，在细胞培养模型中，无法测量细胞培养片的蛋白质渗透性，我们测量了反式内皮电阻。 (TER) 在培养的肺血管内皮细胞片上，我们连续测量了大鼠肺血管内皮细胞汇合时的培养细胞片的TER，并测试了培养细胞片的几种激动剂和拮抗剂。 PKC 激动剂以剂量依赖性方式减少培养的细胞片，而 PKC 激动剂的载体 DMSO 则失败。这些发现也支持了离体大鼠肺实验的结果。肺血管内皮细胞中的肌动蛋白纤维在没有任何刺激的情况下存在于细胞质的周围区域，但是，它们被PKC激动剂解聚并在细胞核周围移动，这些结果表明肺血管内皮细胞发生了变化。当它们受到某种刺激时，PKC会解聚肌动蛋白纤维，从而改变其形态，最终导致肺微血管通透性增加。