The overall objective of the proposed research is to define the secretory functions of normal and acutely injured type II alveolar epithelial cells with central focus on the pathophysiology of lamellar body, the storage organelle of lung surfactant phospholipids and surfactant associated proteins (SAP). The proposed studies will test the hypothesis that lysosomal enzymes modulate SAP concentration within lamellar bodies thereby determining precursor-product relationships between stored, secreted and recycled surfactant. This hypothesis predicts that factors inhibiting the exocrine function of type II cells would result in accelerated proteolysis within the """"""""autophagic lamellar body pool"""""""", selective SAP deficiency and impairment of surfactant recycling, thereby mediating quantitative and compositional alterations of surfactant. Integrated in vivo and in vitro model systems for ozone induced type II cell dysfunction will be used to characterize the subcellular localization of SAP and to define the rate of proteolysis within lamellar bodies as well as the compositional, quantitative relationships between stored and secreted lamellar body constituents including major phospholipids, SAP and lysosomal enzymes. Protein degradation within lamellar bodies from normal and ozone exposed rats will be measured, and the in vitro effects of lysosomal inhibitors on proteolysis and SAP in isolated lamellar bodies will be assessed. Additional in vitro experiments will determine the effects of lysosomal inhibitors and surfactant liposomes, with or without stimulation by Beta-agonists, on the composition of stored and secreted lamellar bodies. Biochemical, immunochemical and lysosomal enzyme assays will be correlated with ultrastructural, immunocytochemical and morphometric data to establish precursor-product relationships between stored, secreted and reutilized lamellar body constituents. The experimental strategy is expected to provide a pathogenetic basis for clinically monitoring the severity of acute type II cell dysfunction and for initiating appropriate therapeutic modalities in adult respiratory distress syndrome.
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