Generation of neutrophil chemoattractants by phagocytosing bovine mammary macrophages

Macrophages from bovine mammary gland were cultured in vitro and the growth medium collected at intervals. Using an in vitro system in which neutrophils migrated under agarose, both chemotactic and chemokinetic activity for bovine neutrophils was detected in supernatants of macrophage cultures to which heat killed preopsonised Staphylococcus aureus had been added. The suspensions of killed bacteria were not themselves chemotactic for neutrophils and no chemotactic activity was present either in supernatants from unstimulated macrophage cultures or in sonicated macrophages. The chemotaxin(s) was generated within two hours of the addition of staphylococci to the cultures and was largely stable to heating at 56 degrees C for 45 minutes, although its activity was reduced by boiling for 15 minutes. Traces of proteolytic activity were also detected in some supernatants. Substantial proteolytic activity was found in lysates of neutrophils. Unlike chemotaxis, proteolytic activity was suppressed by addition of milk from early lactation and containing high natural levels of protease inhibitors. Proteolytic activity was destroyed by boiling for 15 minutes.