The involvement of calcium and membrane fusion events in neutrophil secretion and respiratory burst

Abstract

Neutrophil granulocytes play an important role in host defence during the early stages of microbial infection. The neutrophils have to leave the blood stream and migrate out into the tissue where they are able to phagocytose and kill microbes as well as dispose of cell debris from damaged tissues. Killing methods used by the neutrophils include production of oxygen free radicals, both extracellular release and production at intracellular sites, and release of antimicrobial enzymes from preformed membrane vesicles. These mechanisms work in synergy to create a highly toxic environment for the prey. The efficient removal of pathogens may also have deleterious effects to the host itself, as uncontrolled neutrophil activation may result in damage to host tissues, as seen in various diseases. Increased knowledge of the regulation of neutrophil secretion is thus not only of basic scientific interest but may also have clinical implications. Elevation of the intracellular concentration of free calcium ions ([Ca2+]i) has long been regarded as an important signalling molecule in neutrophil activation. Also, almost all aspects of neutrophil function rely on extensive membrane re-modelling, achieved through membrane fusion events. Neutrophil function was followed through measurements of chemiluminescence (free radical production), Fura-2 fluorescence (intracellular [Ca2+]) and membrane capacitance (secretion).The production of free radicals during phagocytosis was evaluated by use of a technique that minimise the temporal delay in the phagocytic process. An intact cytoskeleton was found to be important for continued production of oxygen radicals. Furthermore, plasma membrane localised calcium channels were found to play crucial roles in the release of radicals as well as in control of granule-to-plasma membrane fusion and subsequent release of granular content. The mere influx of calcium ions does however not seem to be important. Intracellular fusion between different granule types appears to be of importance for processing of the newly produced oxygen radicals, and phospholipase A2 was found to participate in the regulation of this process.