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Cytosolic free calcium elevation mediates the phagosome-lysosome fusion during phagocytosis in human neutrophils

Jaconi, Marisa
Magnusson, K. E.
Sjogren, M.
Stendahl, O.
Published in Journal of Cell Biology. 1990, vol. 110, no. 5, p. 1555-1564
Abstract Cytosolic free calcium ([Ca2+]i) and fusion of secondary granules with the phagosomal membrane (phagosome-lysosome fusion, P-L fusion) were assessed in single adherent human neutrophils during phagocytosis of C3bi-opsonized yeast particles. Neutrophils were loaded with the fluorescent dye fura2/AM and [Ca2+]i was assessed by dual excitation microfluorimetry. Discharge of lactoferrin, a secondary granule marker into the phagosome was verified by immunostaining using standard epifluorescence, confocal laser scanning and electron microscopy. In Ca2(+)-containing medium, upon contact with a yeast particle, a rapid rise in [Ca2+]i was observed, followed by one or more Ca2+ peaks (maximal value 1,586 nM and median duration 145 s): P-L fusion was detected in 80% of the cells after 5-10 min. In Ca2(+)-free medium the amplitude, frequency and duration of the [Ca2+]i transients were decreased (maximal value 368 nM, mostly one single Ca2+ peak and median duration 75 s): P-L fusion was decreased to 52%. Increasing the cytosolic Ca2+ buffering capacity by loading the cells with MAPT/AM led to a dose-dependent inhibition both of [Ca2+]i elevations and P-L fusion. Under conditions where basal [Ca2+]i was reduced to less than 20 nM and intracellular Ca2+ stores were depleted, P-L fusion was drastically inhibited while the cells ingested yeast particles normally. P-L fusion could be restored in Ca2(+)-buffered cells containing ingested particles by elevating [Ca2+]i with the Ca2(+)-ionophore ionomycin. The present findings directly indicate that although the ingestion step of phagocytosis is a Ca2(+)-independent event, [Ca2+]i transients triggered upon contact with opsonized particles are necessary to control the subsequent fusion of secondary granules with the phagosomal membrane.
Keywords Benzofurans/diagnostic useCalcium/metabolism/ physiologyCytosol/metabolismEgtazic Acid/diagnostic useFluorescent Antibody TechniqueFluorescent Dyes/diagnostic useFura-2/ analogs & derivativesHumansImmunohistochemistryLactoferrin/metabolismLysosomes/metabolism/ physiologyMembrane Fusion/physiologyMicroscopy, ElectronNeutrophils/ metabolismPhagocytosis/ physiologyPhagosomes/metabolism/ physiology
PMID: 2110568
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JACONI, Marisa et al. Cytosolic free calcium elevation mediates the phagosome-lysosome fusion during phagocytosis in human neutrophils. In: The Journal of Cell Biology, 1990, vol. 110, n° 5, p. 1555-1564. doi: 10.1083/jcb.110.5.1555 https://archive-ouverte.unige.ch/unige:7350

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Deposited on : 2010-06-21

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