CONFOCAL-MICROSCOPIC AND ELECTRON-MICROSCOPIC LOCALIZATION OF FITC-ALBUMIN IN H2O2-INDUCED PULMONARY-EDEMA

Fluorescein isothiocyanate-conjugated bovine serum albumin (FITC-BSA) was used to detect sites of protein leakage in rat lungs perfused for 15-60 min with H2O2 (75, 150, and 300 mu M) Leaky vessels were localiz ed by confocal laser microscopy. Endothelial routes of protein leakage were identified by electron microscopy after photoconversion of FITC- BSA to an osmiophilic diaminobenzidine product. Transport of FITC-BSA into the alveolar interstitium was assessed by immunogold labeling and anti-FITC antibodies. We detected leakage of FITC-BSA through transen dothelial gaps in the pulmonary arterial endothelium after 30 min of p erfusion with 300 mu M H2O2 and after 60 min of perfusion with 150 mu M H2O2, Junctional permeability and distribution of ZO-1 protein in th e arterial endothelium were unchanged. Microvascular permeability to F ITC-BSA was not increased in lungs perfused with H2O2. In lungs perfus ed with 300 mu M H2O2, progressive extravasation of albumin was associ ated with significant increases in water content and perfusing pressur e. We conclude that the pulmonary arterial endothelium is the primary target of circulating H2O2.