Abstract
Mast cells (MCs) have detrimental functions in the context of numerous pathologies, and regimens aimed at neutralizing MCs or individual MC products can thus be of therapeutic value. One way to target MCs in disease is to selectively induce MC apoptosis, but there is so far no agent available that selectively induces apoptosis in MCs. Mast cells are heavily loaded with secretory granules containing large amounts of fully active proteases bound to serglycin proteoglycan. Damage to the secretory granules will thus lead to the release of serglycin-protease complexes into the cytosol. A potential consequence of this would be that the unleashed granular proteases cause apoptosis by proteolytic activation of proapoptotic compounds located in the cytosol. Indeed, we have recently found that MCs are highly sensitive to apoptosis induced by permeabilization of the secretory granules. In this chapter, we describe the methods used to study MC apoptosis induced by this novel, secretory granule-mediated pathway.
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Abbreviations
- AO:
-
Acridine orange
- CHAPS:
-
3-[(3-Cholamidopropyl)dimethylammonio]-1-propanesulfonate hydrate
- DMSO:
-
Dimethyl sulfoxide
- DTT:
-
Dithiothreitol
- EDTA:
-
Ethylenediaminetetraacetic acid
- EGTA:
-
Ethylene glycol tetraacetic acid
- FITC:
-
Fluorescein isothiocyanate
- HEPES:
-
4-(2-Hydroxyethyl)-1-piperazineethanesulfonic acid
- LDH:
-
Lactate dehydrogenase
- LLME:
-
H-Leu-Leu-OMe⋅HBr
- NAO:
-
Acridine orange 10-nonyl bromide
- PI:
-
Propidium iodide
- PIPES:
-
Piperazine-N,N′-bis(2-ethanesulfonic acid
- R110:
-
Rhodamine 110
- SDS-PAGE:
-
Sodium dodecyl sulfate polyacrylamide gel electrophoresis
- Z-DEVD-FMK:
-
Z-Asp(O-Me)-Glu(O-Me)-Val-Asp(O-Me) fluoromethyl ketone
- Z-VAD-FMK:
-
N-Benzyloxycarbonyl-Val-Ala-Asp(O-Me) fluoromethyl ketone
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Acknowledgments
The authors of this article receive support from The Swedish Research Council, Formas, King Gustaf V 80-year Anniversary Fund, Torsten and Ragnar Söderberg Foundation, The Vårdal Foundation, The Swedish Society of Medicine, Åke Wiberg Foundation, Konsul Th C Bergh Foundation, and The Swedish Cancer Foundation.
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Melo, F.R., Wernersson, S., Pejler, G. (2015). Induction of Mast Cell Apoptosis by a Novel Secretory Granule-Mediated Pathway. In: Hughes, M., McNagny, K. (eds) Mast Cells. Methods in Molecular Biology, vol 1220. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1568-2_20
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DOI: https://doi.org/10.1007/978-1-4939-1568-2_20
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Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-1567-5
Online ISBN: 978-1-4939-1568-2
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