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.
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