Resolution of acute lung injury (ALl) is dependent on temporally and spatially regulated elimination of both inflammatory cells and mesenchymal cells from the alveolar space. The origin of mesenchymal cells and their mechanisms of activation and clearance in ALI/ARDS are unclear. We have isolated CD45(-) alveolar mesenchymal cells (AMCs) expressing fibroblast/myofibroblast markers (collagen I, prolyl-4- hydroxylase, alpha-smooth muscle actin) from bronchoalveolar lavage (BAL) of patients with ALl. AMCs proliferate in vitro and possess distinct, yet stable cellular phenotypes. AMCs from patients with fibroproliferative ALl vs resolving ALl are 'intrinsically' resistant to apoptosis with constitutive activation of the pro-survival PI3K-Akt pathway. This activated AMC phenotype may be mediated by an autocrine TGF-beta1/thrombospondin-1 activation loop. Hyperoxia may further promote the anti-apoptotic phenotype by activating the PI3K-Akt pathway in AMCs, in contrast to primarily pro-apoptotic signaling in alveolar epithelial cells. Thus, 'extrinsic' alveolar microenvironmental factors and 'intrinsic' factors modulate AMC phenotype and contribute to dysregulated lung repair by promoting the activation and accumulation of AMCs within alveolar spaces.HYPOTHESIS: Intra-alveolar accumulation of AMCs with an activated, apoptosis-resistant phenotype is predictive of dysregulated lung repair and persistent ALl. Both 'intrinsic' and 'extrinsic' (alveolar microenvironment) factors regulate this AMC phenotype.
SPECIFIC AIMS are to: (1) determine whether an 'intrinsically' autocrine TGF-beta1/TSP-1 activation loop regulates the anti-apoptotic AMC phenotype in patients with persistent ALI/ARDS, (2) determine if elevated levels of activated TGF-beta1 and hyperoxia (alveolar microenvironmenta factors) promote an anti-apoptotic phenotype of AMCs, (3) prospectively determine whether the number of AMCs (CD45-/PH+/o_-SMA+ cells recovered by BAL), their phenotype (apoptosis, proliferation, oxidative burst activity), and/or activated TGF- 131in the alveolar microenvironment is predictive of persistent ALl and clinical outcomes in a cohort of ALl patients and (4) determine if the presence of TGF-131 gene polymorphisms (at codon 10 and/or 25) is predictive of persistent/fibroproliferative ALI.
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