MAPK signaling in injury, proliferation and fibrosis

天冬氨酸在体外和体内激活肺泡II型上皮（C10）细胞中的丝裂原活化蛋白激酶（MAPK）级联。在C10细胞中，这些信号传导事件先于fos/jun家族成员的mRNA水平的早期增加和fra-1和Fra-1/Jun（AP-1）复合物的mRNA水平的更持久的增加。我们推测，石棉激活ERK和JNK通路导致AP-1家族成员和亚基组成的表达变化，然后支配细胞周期的变化，损伤和随后的肺上皮细胞增殖。在具体目标#1中，我们将使用转染技术在C10肺泡II型上皮细胞中确定石棉对ERK和JNK级联的激活是否与fos/jun成员、c-fos、c-jun和fra-1的表达增加、AP-1复合物的组成、AP-1依赖性基因表达的反式激活以及随后的结果（细胞周期改变、损伤和代偿性增殖）有因果关系。在具体目标#2中，我们将使用过表达显性阴性（dn）MEK 1或dnJNK 1与肺上皮细胞启动子（CC 10和SPC）的转基因小鼠评估ERK和JNK通路是否与石棉肺小鼠模型中远端细支气管和肺泡II型上皮细胞的增殖和纤维化的发展存在因果关系。 在具体目标#3中，我们将使用激光捕获显微切割和真实的时间PCR（TaqMan）确定暴露于石棉的小鼠的远端细支气管和肺泡II型上皮细胞中c-fos、c-jun和fra-1表达是否增加，以及在表达上皮细胞特异性dnMEK 1、dnJNK 1的转基因小鼠或这些小鼠的回交中模式是否改变以阻断两种途径。该项目的目标与该项目的主题有关，即MAPK级联在上皮细胞损伤和增殖中的作用，并将确定这些细胞信号传导事件的修饰是否会在体外和石棉肺吸入模型中改变这些结果。我们的假设是新颖的，因为它们将提供一个机制框架MAPK途径和AP-1亚基组成的原因上皮细胞损伤和代偿性增生。

Asbestos activates mitogen-activated protein kinase (MAPK) cascades in alveolar type II epithelial (C10) cells in vitro and in vivo. In C10 cells, these signaling events precede early increases in mRNA levels of fos/jun family members and more protracted increases in mRNA levels of fra-1 and Fra-1/Jun (AP-1) complexes. We hypothesize that activation of ERK and JNK pathways by asbestos leads to changes in expression of AP-1 family members and subunit composition that then govern cell cycle changes, injury and subsequent proliferation of pulmonary epithelial cells. In Specific Aim #1, we will determine in C10 alveolar type II epithelial cells using transfection techniques whether activation of ERK and JNK cascades by asbestos are linked causally to increased expression of the fos/jun members, c-fos, c-jun and fra-1, composition of AP-1 complexes, transactivation of AP-1 dependent gene expression, and subsequent outcomes (cell cycle alterations, injury and compensatory proliferation). In Specific Aim #2, we will assess using transgenic mice over-expressing dominant negative (dn)MEK1 or dnJNK1 with lung epithelial cell promoters (CC10 and SPC), whether ERK and JNK pathways are causally related to proliferation of distal bronchiolar and alveolar type II epithelial cells and the development of fibrosis in a murine model of asbestosis. In Specific Aim #3, we will determine using laser capture microdissection and real time PCR (TaqMan) whether c-fos, c-jun and fra-1 expression is increased in distal bronchiolar and alveolar type II epithelial cells of mice exposed to asbestos, and whether patterns are modified in transgenic mice expressing epithelial cell-specific dnMEK1, dnJNK1 or backcrosses of these mice to block both pathways. The goals of this Project are related to the theme of the Program Project, the role of MAPK cascades in epithelial cell injury and proliferation, and will ascertain whether modification of these cell signaling events modifies these outcomes in vitro and in an inhalation model of asbestosis. Our hypothesis are novel in that they will provide a mechanistic framework for MAPK pathways and AP-1 subunit composition in the causation of epithelial cell injury and compensatory hyperplasia.