Sequential Degranulation of the Two Types of Polymorphonuclear Leukocyte Granules During Phagocytosis of Microorganisms

Abstract

The sequential discharge of neutrophilic polymorphonuclear leukocyte (PMN) granules—azurophils and specifics—was investigated by electron microscopy and cytochemistry. Thus the enzyme content of PMN phagocytic vacuoles was determined at brief intervals after phagocytosis of bacteria, utilizing peroxidase as a marker enzyme for azurophil granules, and alkaline phosphatase for specifics. At 30 s, approximately half the phagocytic vacuoles were reactive for alkaline phosphatase, whereas none contained peroxidase. Peroxidase-containing vacuoles were rarely seen at 1 min, but by 3 min, vacuoles containing both enzymes were consistently present. Alkaline phosphatase was found in both small and large vacuoles, whereas peroxidase was visible only in large ones. By 10 min, very big phagocytic vacuoles containing considerable amounts of reaction product for both enzymes were evident. These observations indicate that the two types of PMN granules discharge in a sequential manner, specific granules fusing with the vacuole before azurophils. In an earlier paper, we reported that the pH of phagocytic vacuoles drops to 6.5 within 3 min and to ∼4 within 7–15 min. Substances known to be present in specific granules (alkaline phosphatase, lysozyme, and lactoferrin) function best at neutral or alkaline pH, whereas most of those contained in azurophil granules (i.e., peroxidase and the lysosomal enzymes) have pH optima in the acid range. Hence the sequence of granule discharge roughly parallels the change in pH, thereby providing optimal conditions for coordinated activity of granule contents.

The bovine neutrophil: Structure and function in blood and milk

“…The inflammatory mediators PAF and tumor necrosis factor (TNF) induce an increase in size of bovine PMN [108], slow diapedesis of PMN through the microvasculature of the mammary gland, and regulate movement of PMN to the infection site. Alkaline phosphatase is also present in secondary granules [13].…”

“…Milk PMN have reduced numbers of secondary and "novel" granules compared to blood PMN. Secondary and "novel" granules are the first granules to be released into phagosomes and are extruded from PMN in greater number during diapedesis into milk than are azurophilic granules [13,83]. Glycogen particles are also present throughout the cytoplasm of milk PMN.…”

“…Like other species, bovine PMN contain azurophilic and specific (secondary) granules. Azurophilic or primary granule is the first granule to appear during the progranulocyte stage of granulopoiesis [13,14,83]. They are large electron-dense granules.…”

Phagocytosis of zymosan by human neutrophils

“…The sequential events in phagocytosis including particle recognition, attachment, engulfment and degranulation have been well studied by light and electron microscopy by numerous investigators (1,5,7,8,10,22). Phagocytosis of foreign particles is usually followed by lysis and degradation of the microorganisms through the action of hydrolases and antibacterial proteins contained within cytoplasmic granules (1,8,10,17,18,22).…”

“…The sequential events in phagocytosis including particle recognition, attachment, engulfment and degranulation have been well studied by light and electron microscopy by numerous investigators (1,5,7,8,10,22). Phagocytosis of foreign particles is usually followed by lysis and degradation of the microorganisms through the action of hydrolases and antibacterial proteins contained within cytoplasmic granules (1,8,10,17,18,22). Human Abbreviations: LF = lactoferrin; MPO ffi myeloperoxidasr SEM = scanning electron microscopy; TEM = transmission electron microscopy Springer-Verlag 0105-1938/84/0049/0315/$01.60 neutrophils contain two distinct types of granules which have been analyzed morphologically and biochemically (2,3,4,5,6,16,17,21).…”