Reassessment of the microbicidal activity of reactive oxygen species and hypochlorous acid with reference to the phagocytic vacuole of the neutrophil granulocyte

During phagocytosis, neutrophils undergo a burst of respiration in which oxygen is reduced to superoxide (O-2(-)), which dismutates to form H2O2. Myeloperoxidase (MPO) is discharged from the cytoplasmic granules into the phagosome following particle ingestion. It is thought to utilize H2O2 to oxidize halides, which then react with and kill ingested microbes. Recent studies have provided new information as to the concentration of O-2(-) and proteins, and the pH, within the vacuole. This study was conducted to examine the antimicrobial effect of O-2(-), H2O2 and hypochlorous acid under these conditions and it was found that the previously described bactericidal effect of these agents was reversed in the presence of granule proteins or MPO. To establish which cellular proteins were iodinated by MPO, cellular proteins and bacterial proteins, iodinated in neutrophils phagocytosing bacteria in the presence of I-125, were separated by 2D gel electrophoresis. Iodinated spots were detected by autoradiography and the oxidized proteins were identified by MS. The targets of these iodination reactions were largely those of the host cell rather than those of the engulfed microbe.