Hydrogen peroxide-scavenging properties of sheep airway mucus

Reactive oxygen species released from luminal phagocytes in the airway can potentially injure the airway epithelium. Naturally occurring oxygen radical scavengers must therefore exist to protect the epithelium. This study was designed to determine whether the high-molecular-weight fraction of normal sheep tracheal mucus has hydrogen peroxide (H2O2)-scavenging activity. Lyophilized mucus from 10 sheep was reconstituted in phosphate-buffered saline (PBS) or Krebs-Henseleit buffer. H2O2 was added to these mucus samples to a final concentration of 15 microM, and the level of H2O2 remaining was measured over a 10 min period. From a zero-time level of 17 +/- 1.8 microM (mean +/- SD), the H2O2 concentration fell within 10 min to 8 +/- 1.7 microM in 0.05%; to 3.9 +/- 2.2 microM in 0.1%; to 2.6 +/- 2.4 microM in 0.2%; and to 1.2 +/- 1.5 microM in 0.4% mucus reconstituted in PBS. The results obtained in Krebs-Henseleit buffer were similar. The disappearance of H2O2 was not due to the transformation into hydroxyl radicals. Heat and acid denaturation and cleavage of carbohydrate-free peptides from glycoproteins by pronase E treatment abolished the scavenging potential. Fractionation of 0.4% mucus samples according to molecular weight by gel filtration revealed that only one fraction with proteins of M(r) > 110 kD contained the active scavenger. Polyacrylamide gel electrophoresis and lectin blotting with Ulex europaeus I (UEAI) showed that both the whole mucus and the actively scavenging gel filtration fraction contained a glycoprotein that comigrated with a 205 kD molecular weight marker.(ABSTRACT TRUNCATED AT 250 WORDS)