PERTICIPATION OF INTRACELLULAR SIGNAL TRANSDUCTION PATHWAY AND RE-PERFUSION LUNG INJURY AFTER LUNG TRANSPLANTATION

结果表明，中性粒细胞粘附内皮细胞后，通过黄嘌呤氧化酶（XO）产生活性氧诱导了中性粒细胞诱导的肺损伤；通常，XO不存在于中性粒细胞中，而存在于内皮细胞中。因此，我们推测这种肺损伤是否由内皮细胞自身的活化引起。在离体灌注大鼠肺实验中，我们使用蛋白激酶C拮抗剂研究了细胞内信号转导系统是否参与了活化中性粒细胞引起的肺血管内皮细胞损伤。我们发现PKC拮抗剂以剂量依赖的方式改善了活化中性粒细胞引起的肺血管损伤。此外，我们发现不含中性粒细胞的PKC激动剂可增加肺血管通透性。另一方面，在细胞培养模型中，不可能测量细胞培养片的蛋白质通透性，我们测量了培养的肺血管内皮细胞片的跨膜上皮电阻（TER）。高渗透压暴露的主要影响是TER的增加，表明诱导屏障加强。高渗透压暴露也增加了细胞周围黏附激酶的活性和e -钙粘蛋白的积累。同时，肌动蛋白丝密度显著增加。蛋白激酶C的激动剂PMA（肉豆蔻酸磷酯）降低了TER，表明肌动蛋白细丝的解聚。另一方面，肌动蛋白丝的解聚合剂细胞松弛素D也降低了TER，表明参与了肌动蛋白的解聚合。肌动蛋白对高渗暴露的反应增强，但对TER的反应不受影响，表明内皮屏障增强是肌动蛋白独立发生的。然而，在表达局灶黏附激酶激酶依赖突变体的单层细胞中，高渗透压诱导的局灶黏附激酶活性增加和外周E-cadherin增强被阻断，诱导的TER增加明显减弱。这些结果表明，EC暴露于高渗透压刺激下，局灶黏附激酶的参与是建立屏障强化的关键。少

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. Besides, 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-en … More dothelial electrical resistance (TER) across the cultured pulmonary vascular endothelial cell sheets. The dominanat effect of hyperosmolar exposure was an increase in the TER, indicating the induction of barrier strengthening. Hyperosmolar exposure also increased activity of focal adhesion kinase and E-cadherin accumulation at the cell periphery. Concomitantly, the density of actin filaments increased markedly. PMA (phobor myristate acetate), an agonist of Protein kinase C, decreased the TER, indicating a de-polymerization of actin filaments. On the other hand, cytochalasin D, de-polymerize agents for actin filaments, also decreased TER, indicating participation of actin de-polymerization. The actin response to hyperosmolar exposure was enhanced although the response in TER was unaffected, indicating that the endothelial barrier enhancement occurred independently actin. However, in monolayers expressing a kinase-dependent mutant of focal adhesion kinase, the hyperosmolarity-induced increase in activity of focal adhesion kinase and peripheral E-cadherin enhancement were blocked and the induced increase of TER was markedly blunted. These findings indicate that EC exposed to hyperosmolar challenge, the involvement of focal adhesion kinase was critical in establishing barrier strengthening. Less