Imaging bioluminescent indicators shows Ca2+ and ATP permeability thresholds in live cells attacked by complement

Immunology
G B Sala-NewbyA K Campbell

Abstract

A series of permeability thresholds to Ca2+ metabolites and macromolecules, occurring at different times when cells are attacked by complement, has been established by imaging HeLa cells transiently expressing a recombinant cytosolic fusion protein of firefly luciferase and aequorin (luciferase-aequorin) to measure changes in ATP and cytosolic free Ca2+. Nuclear fluorescence of propidium was used as a measure of permeability to small molecules, and luciferase activity imaged to assess lysis. The rise in cytosolic free Ca2+ observed after C9 attack preceded by at least 60 s both the increase in propidium fluorescence, measured in single cells, and the decrease in ATP monitored by luciferase light emission. These effects were dependent on the concentration of C9. At concentrations of C9 up to 4 micrograms/ml no loss of luciferase-aequorin protein was detected at the end of the experiment. Thus the membrane integrity of the cells remained intact, even though the cells were permeable to propidium. These results confirmed our earlier observations that propidium permeability in cells attacked by complement was not a reliable measure of cell death. They also show that it is vital to take account of cellular heterogeneity if the mechan...Continue Reading

References

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Citations

Aug 30, 2007·Science Progress·Lubov Y Brovko, Mansel W Griffiths
Sep 24, 2013·Photochemical & Photobiological Sciences : Official Journal of the European Photochemistry Association and the European Society for Photobiology·Mikhail I Koksharov, Natalia N Ugarova
Aug 26, 2020·British Journal of Pharmacology·Jubayer RahmanClaudia Kemper
Jan 30, 2002·Luminescence : the Journal of Biological and Chemical Luminescence·Lee F Greer, Aladar A Szalay
Jul 16, 2002·Annual Review of Biomedical Engineering·Christopher H Contag, Michael H Bachmann

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