Applying label-free dynamic mass redistribution assay for studying endogenous FPR1 receptor signalling in human neutrophils

Hanna B Christensen; David E Gloriam; Daniel Sejer Pedersen; Jack B Cowland; Niels Borregaard; Hans Bräuner-Osborne

doi:10.1016/j.vascn.2017.07.003

Applying label-free dynamic mass redistribution assay for studying endogenous FPR1 receptor signalling in human neutrophils

Hanna B Christensen, David E Gloriam, Daniel Sejer Pedersen, Jack B Cowland, Niels Borregaard, Hans Bräuner-Osborne

Department of Clinical Medicine

7 Citations (Scopus)

Abstract

INTRODUCTION: The label-free dynamic mass redistribution-based assay (DMR) is a powerful method for studying signalling pathways of G protein-coupled receptors (GPCRs). Herein we present the label-free DMR assay as a robust readout for pharmacological characterization of formyl peptide receptors (FPRs) in human neutrophils.

METHODS: Neutrophils were isolated from fresh human blood and their responses to FPR1 and FPR2 agonists, i.e. compound 43, fMLF and WKYMVm were measured in a label-free DMR assay using Epic Benchtop System from Corning®. Obtained DMR traces were used to calculate agonist potencies.

RESULTS: The potencies (pEC50) of fMLF, WKYMVm and compound 43, determined on human neutrophils using the label-free DMR assay were 8.63, 7.76 and 5.92, respectively. The DMR response to fMLF, but not WKYMVm and compound 43 could be blocked by the FPR1-specific antagonist cyclosporin H.

DISCUSSION: We conclude that the DMR assay can be used, and complements more traditional methods, to study the signalling and pharmacology of endogenous FPR receptors in human neutrophils.

Original language	English
Journal	Journal of Pharmacological and Toxicological Methods
Volume	88
Issue number	Pt 1
Pages (from-to)	72-78
Number of pages	7
ISSN	1056-8719
DOIs	https://doi.org/10.1016/j.vascn.2017.07.003
Publication status	Published - Nov 2017

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Applying label-free dynamic mass redistribution assay for studying endogenous FPR1 receptor signalling in human neutrophils. / Christensen, Hanna B; Gloriam, David E; Pedersen, Daniel Sejer et al.

In: Journal of Pharmacological and Toxicological Methods, Vol. 88, No. Pt 1, 11.2017, p. 72-78.

Research output: Contribution to journal › Journal article › Research › peer-review

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abstract = "INTRODUCTION: The label-free dynamic mass redistribution-based assay (DMR) is a powerful method for studying signalling pathways of G protein-coupled receptors (GPCRs). Herein we present the label-free DMR assay as a robust readout for pharmacological characterization of formyl peptide receptors (FPRs) in human neutrophils.METHODS: Neutrophils were isolated from fresh human blood and their responses to FPR1 and FPR2 agonists, i.e. compound 43, fMLF and WKYMVm were measured in a label-free DMR assay using Epic Benchtop System from Corning{\textregistered}. Obtained DMR traces were used to calculate agonist potencies.RESULTS: The potencies (pEC50) of fMLF, WKYMVm and compound 43, determined on human neutrophils using the label-free DMR assay were 8.63, 7.76 and 5.92, respectively. The DMR response to fMLF, but not WKYMVm and compound 43 could be blocked by the FPR1-specific antagonist cyclosporin H.DISCUSSION: We conclude that the DMR assay can be used, and complements more traditional methods, to study the signalling and pharmacology of endogenous FPR receptors in human neutrophils.",

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T1 - Applying label-free dynamic mass redistribution assay for studying endogenous FPR1 receptor signalling in human neutrophils

AU - Christensen, Hanna B

AU - Gloriam, David E

AU - Pedersen, Daniel Sejer

AU - Cowland, Jack B

AU - Borregaard, Niels

AU - Bräuner-Osborne, Hans

PY - 2017/11

Y1 - 2017/11

N2 - INTRODUCTION: The label-free dynamic mass redistribution-based assay (DMR) is a powerful method for studying signalling pathways of G protein-coupled receptors (GPCRs). Herein we present the label-free DMR assay as a robust readout for pharmacological characterization of formyl peptide receptors (FPRs) in human neutrophils.METHODS: Neutrophils were isolated from fresh human blood and their responses to FPR1 and FPR2 agonists, i.e. compound 43, fMLF and WKYMVm were measured in a label-free DMR assay using Epic Benchtop System from Corning®. Obtained DMR traces were used to calculate agonist potencies.RESULTS: The potencies (pEC50) of fMLF, WKYMVm and compound 43, determined on human neutrophils using the label-free DMR assay were 8.63, 7.76 and 5.92, respectively. The DMR response to fMLF, but not WKYMVm and compound 43 could be blocked by the FPR1-specific antagonist cyclosporin H.DISCUSSION: We conclude that the DMR assay can be used, and complements more traditional methods, to study the signalling and pharmacology of endogenous FPR receptors in human neutrophils.

AB - INTRODUCTION: The label-free dynamic mass redistribution-based assay (DMR) is a powerful method for studying signalling pathways of G protein-coupled receptors (GPCRs). Herein we present the label-free DMR assay as a robust readout for pharmacological characterization of formyl peptide receptors (FPRs) in human neutrophils.METHODS: Neutrophils were isolated from fresh human blood and their responses to FPR1 and FPR2 agonists, i.e. compound 43, fMLF and WKYMVm were measured in a label-free DMR assay using Epic Benchtop System from Corning®. Obtained DMR traces were used to calculate agonist potencies.RESULTS: The potencies (pEC50) of fMLF, WKYMVm and compound 43, determined on human neutrophils using the label-free DMR assay were 8.63, 7.76 and 5.92, respectively. The DMR response to fMLF, but not WKYMVm and compound 43 could be blocked by the FPR1-specific antagonist cyclosporin H.DISCUSSION: We conclude that the DMR assay can be used, and complements more traditional methods, to study the signalling and pharmacology of endogenous FPR receptors in human neutrophils.

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SN - 1056-8719

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JO - Journal of Pharmacological and Toxicological Methods

JF - Journal of Pharmacological and Toxicological Methods

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Applying label-free dynamic mass redistribution assay for studying endogenous FPR1 receptor signalling in human neutrophils

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