The normal alveolar epithelium develops rapidly during the early postnatal period. Currently little is known about the adaptations of alveolar epithelial cells to injury in either the adult or especially the neonatal lung. Several functional membrane proteins are quantitative markers of alveolar differentiation: Na,K-ATPase for type II cells and gamma-glutamyl transpeptidase (GGT) for type I cells. The sodium pump is partially responsible for clearance of salt and water from the alveolar space. GGT may play a role in glutathione recycling and therefore may be important in oxidant stress. Hyperoxia is a representative oxidant stress that is a model of inflammatory lung damage. Also, high concentrations of oxygen often are used clinically to support infants with damaged lungs. In the adult lung, hyperoxia causes necrosis of type I cells and proliferation of type II cells. There is an early increase in the mRNA levels of Na,K- ATPase in type II cells, prior to their proliferation, that may help defend against pulmonary edema. We believe that neonatal hyperoxic lung injury will disrupt the normal sequence of developmental changes in these proteins accompanying alveolar differentiation and that expression of first Na,K- ATPase and then GGT will be upregulated during the neonatal lung injury and repair. We hypothesize that this regulation will occur at the level of the cellular mRNA levels. We propose to examine changes in these functional membrane proteins during normal perinatal rat lung development and during hyperoxic injury and repair in the early postnatal period. Specifically changes in Na,K-ATPase and GGT localization (proteins by immunofluorescence and immunoEM; mRNA by in situ hybridization), quantity (Western blotting), activity, and mRNA levels (Northern blotting) will be characterized during normal development. Then the changes occurring during postnatal hyperoxic injury will be determined and contrasted with the normal sequence. In vitro hyperoxic injury model of cultured neonatal lung cells will be used to confirm which cell type(s) are involved. These experiments should provide insight into the effects of injury on the normal development of the alveolar epithelium and on the regulation of proteins responsible for several specialized functions of type I and II cells.
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