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* From the Departments of Medicine and Pediatrics, University of Minnesota, Minneapolis, MN, and Department of Medicine, Washington University, St. Louis, MO. Supported by NIH SCOR in Acute Lung Injury HL50152 and an ALA Career Investigator Award.
Correspondence to: David H. Ingbar, MD, FCCP, University of Minnesota School of Medicine, 420 Delaware St SE, Box 276, Minneapolis, MN 55455
Hyperoxia causes
lung injury due to increased oxidants and inflammation. We sought to
assess whether oxidative modification of proteins occurred in this
model of rat acute lung injury and to determine if specific membrane
transport proteins were affected. Using antibodies to hypochlorous
acid-modified low-density lipoprotein and tyrosine
groups, western blotting revealed marked increases in specific protein
bands at approximately 100 and 60 kd in rat lungs at 60 h of
hyperoxic exposure. Using the former antibody for immunocytochemistry,
there was little reactivity in normal lungs, but hyperoxia induced
striking staining of airway and alveolar epithelium that persisted with
only moderate decreases through 7 days of recovery. Since in prior data
hyperoxia decreased the maximal enzyme velocity of type II cell
Na,K-adenosine triphosphatase, we hypothesized that this was due to
pump oxidation and assessed whether nitrotyrosine residues were present
on this integral membrane protein from normoxic, hyperoxic, and
recovering lungs. Using immunoprecipitation and then western blotting,
the ß1-subunit, but not the
1-subunit, had
detectable nitrotyrosine even in normoxic lung. Hyperoxia led to a
statistically significant, but small (approximately 30%), increase in
nitrotyrosine of the sodium pump ß1-subunit. This
decreased rapidly during the recovery phase. These data indicate that
during in vivo hyperoxia, newer oxidant species cause
significant protein modification and suggest that important epithelial
cell functional proteins can be
affected.
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