Mobilization and exocytosis of specific (secondary) granules by human neutrophils during adherence to nylon wood in filtration leukapheresis (FL)

Abstract

We studied the interaction of human neutrophils with nylon wool in vitro and during filtration leukapheresis (FL) to better understand cellular events detrimental to neutrophils that occur during FL. When neutrophils purified from blood by gradient centrifugation and sedimentation were incubated with nylon wool in vitro, release of lysozyme from the cells occurred rapidly and to a degree that was related to the ratio of nylon wool bulk to the number of cells. Extracellular release of lysozyme was greater than that of β-glucuronidase, and this differential release of granule-associated enzymes reflected a preferential degranulation of the neutrophil-specific (or secondary) granules. Specific and azurophil (primary) granules were separated from neutrophils purified directly from blood and also from cells obtained by FL, and a partial degranulation of specific granules with loss of their associated lysozyme was found with FL neutrophils. Exposure of neutrophils to nylon wool in vitro was found to cause extracellular release of a granule product that activates serum complement and generates the chemotactic C5 fragment, C5a. Plasma from blood leaving nylon wool filters during FL was found to have increased lysozyme activity, reduced hemolytic complement, and increased chemotactic activity attributable in part to C5a production. In addition, neutrophils obtained by FL were found to have changes in surface charge similar to those caused by the exposure of neutrophils to chemotactic factors (such as C5a) or to degranulating stimuli. When plasma and leukocyte-enriched blood were diverted by a cell separator before passage through nylon wool filters during FL, complement activation and consumption were evident only in plasma from blood containing leukocytes, indicating that extracorporeal complement activation during FL is dependent in large part upon the interaction of leukocytes with nylon wool. Detectable quantities of histamine were not released within filters during FL, indicating that extracorporeal basophil degranulation with histamine secretion does not occur in FL to a significant degree as does neutrophil degranulation. These studies support the concept that neutrophil degranulation and secretion of granule contents are of central importance to the functional changes that occur in these cells during FL.