ACTIVATED NEUTROPHILS SECRETE STORED ALPHA 1-ANTITRYPSIN

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Neutrophil elastase (NE), a potent serine protease, is stored in primary granules of neutrophils and released following neutrophil activation. Alpha-1-antitrypsin (alpha1AT), the major inhibitor of NE, is synthesized by mature neutrophils. In the context of the maintenance of tissue homeostasis, we hypothesized that neutrophils may be able to store alpha1AT, thus having it available for release concordantly with NE. Immunofluorescence and quantitative flow cytometry studies of neutrophils and monocytes labeled with fluorescein conjugated alpha1AT-antibody demonstrated larger amounts of cytoplasmic alpha1AT in neutrophils than in monocytes. Study with radioactively labeled methionine and anti-alpha1AT immunoprecipitation analysis showed that, while both neutrophils and monocytes synthesize alpha1AT, the proportion of newly synthesized intracellular alpha1AT was much higher in neutrophils than in monocytes. Flow cytometric analysis showed that in the presence of surface stimulation with cytochalasin B followed by N-formyl-methionyl-leucyl-phenylalanine (fMLP), mean intracellular alpha1AT was decreased in stimulated neutrophils compared with that in resting cells, suggesting that the stored alpha1AT was rapidly released following surface triggering. Evaluation of surface stimulated neutrophils by radioactive methionine labeling and anti-alpha1AT immunoprecipitation demonstrated increased secretion of alpha1AT compared to that of resting neutrophils, with some of the secreted alpha1AT capable of forming complexes with NE. Thus, neutrophils respond to surface stimulation by secreting not only NE but also its inhibitor, alpha1AT. This suggests that these cells have an inherent mechanism for dampening the local effects of NE, its most powerful proteolytic enzyme.