A novel type of cytoplasmic granule in bovine neutrophils.

Open Access
Publisher Website
Google Scholar
Abstract
Cell preparations were obtained containing > 95% neutrophils from feshly drawn bovine blood. The cells were suspended in sucrose and disrupted in a Dounce homogenizer, and the postnuclear supernate was fractionated by zonal differential sedimentation and by isopycnic equilibration. The subcellular fractions were characterized biochemically by testing for marker enzymes and other constituents known to occur in azurophil and specific granules of other species, and by electrophoretic analysis of extracts of the particulate material. In addition, each fraction was examined by random-sampling EM. Bovine neutrophils contained in addition to azurophil and specific granules a 3rd type of granule, not known to occur in neutrophils of other species. These novel granules are larger, denser, and considerably more numerous than the 2 other types. Except for lactoferrin, they lack the characteristic constituents of azurophil granules (peroxidase, acid hydrolases, and neutral proteinases) and of specific granules (vitamin B12-binding protein). Instead, they contain a group of highly cationic proteins not found in the other granules, and they are the exclusive stores of powerful O2-independent bactericidal agents. The fate of the large granules was studied in bovine neutrophils exposed to opsonized particles, the ionophore A 23187 [calcimycin] or phorbol myristate acetate. The appearance in the cell-free media of antibacterial activity and of the characteristic highly cationic proteins as revealed by electrophoresis was monitored and compared with the release of azurophil and specific granule markers. In addition, changes of the relative size of the large granule compartment induced by phagocytosis were assessed by morphometry. Exocytosis of the large granules occurs following both phagocytosis and exposure to soluble stimuli. Like the specific granules, the large granules appear to be discharged by true secretion under conditions where the azurophil granules are fully retained.