Label‐Free Dynamic Mass Redistribution and Bio‐Impedance Methods for Drug Discovery

Grundmann, Manuel

doi:10.1002/cpph.24

Cited by 10 publications

(8 citation statements)

References 56 publications

(89 reference statements)

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“…Dynamic mass redistribution (DMR) measurements were performed in suspension mode as previously described in detail 12 . Briefly, 60,000 freshly isolated neutrophils per well were seeded into a 384‐well biosensor plate in HBSS buffer containing 20 mM HEPES, centrifuged for 30 s at 150 × g and incubated at 28°C for 2 h in the DMR reader before the measurement.…”

Section: Methodsmentioning

confidence: 99%

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Allosteric targeting of the FFA2 receptor (GPR43) restores responsiveness of desensitized human neutrophils

Frei

Nordlohne

Hüser

et al. 2020

Journal of Leukocyte Biology

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The G protein-coupled free fatty acid receptor 2 (FFA2R) is highly expressed on neutrophils and was previously described to regulate neutrophil activation. Allosteric targeting of G proteincoupled receptors (GPCRs) is increasingly explored to create distinct pharmacology compared to endogenous, orthosteric ligands. The consequence of allosteric versus orthosteric FFA2R activation for neutrophil response, however, is currently largely elusive. Here, different FFA2R desensitization profiles in human neutrophils following allosteric or orthosteric activation are reported. Using a set of neutrophil functional assays to measure calcium flux, pERK1/2, chemotaxis, cellular degranulation, and oxidative burst together with holistic and pathway-unbiased whole cell sensing based on dynamic mass redistribution, it is found that the synthetic positive allosteric modulator agonist 4-CMTB potently activates neutrophils and simultaneously alters FFA2R responsiveness toward the endogenous, orthosteric agonist propionic acid (C3) after homologous and heterologous receptor desensitization. Stimulation with C3 or the hierarchically superior chemokine receptor activator IL-8 led to strong FFA2R desensitization and rendered neutrophils unresponsive toward repeated stimulation with C3. In contrast, stimulation with allosteric 4-CMTB engaged a distinct composition of signaling pathways as compared to orthosteric receptor activation and was able to activate neutrophils that underwent homologous and heterologous desensitization with C3 and IL-8, respectively. Moreover, allosteric FFA2R activation could re-sensitize FFA2 toward the endogenous agonist C3 after homologous and heterologous desensitization. Given the fact that receptor desensitization is critical in neutrophils to sense and adapt to their current environment, these findings are expected to be useful for the discovery of novel pharmacological mechanisms to modulate neutrophil responsiveness therapeutically.

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Section: Methodsmentioning

confidence: 99%

“…Dynamic mass redistribution (DMR) measurements were performed in suspension mode as previously described in detail. 12…”

Section: Dynamic Mass Redistribution Measurementsmentioning

confidence: 99%

Allosteric targeting of the FFA2 receptor (GPR43) restores responsiveness of desensitized human neutrophils

Frei

Nordlohne

Hüser

et al. 2020

Journal of Leukocyte Biology

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“…After the cells initially adsorb to the surface, internal signal cascades initiate the attachment of the cells to their environment, followed by the cytoskeleton remodelling leading to the spreading of the cells at the surface. These morphological differences cause changes in the surface area covered by the cell, and in the refractive index of the cell cytoplasm [36,37]. This, in turn, influences the optical path of the captured reflected light in SCORE.…”

Section: Monitoring Morphological Changes Of Cellsmentioning

confidence: 99%

Parallelized label-free monitoring of cell adhesion on extracellular matrix proteins measured by single colour reflectometry

et al. 2021

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The understanding of the initial cell adhesion to biomaterials is crucial for the survival of implants. The manifold possibilities to tailor an implant surface and the diverse requirements for different implant applications necessitate a timesaving and highly parallelized analytical methodology. Due to its intrinsic advantages (label-free, time-resolved, robust against temperature fluctuations, and particularly the multiplexing possibilities), single colour reflectometry (SCORE) is used for the first time to investigate cell adhesion to different extracellular matrix protein–coated surfaces. The excellent correlation between the novel SCORE technology and well-established reference methods proves that the results obtained by using this direct optical method are able to reflect the cell binding processes at the transducer surface. Additionally, the high time resolution of SCORE revealed the differences in the adhesion behaviour of the cells on the different extracellular matrix protein–coated glass slides during the initial adsorption phase and during the spreading of the cells on the surfaces. Therefore, we conclude that SCORE is a perfectly suited methodology for studying the entire cell adsorption process, including morphological changes, and shows great potential for other cell-based sensing applications. Graphical abstract

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“…Dynamic mass redistribution (DMR) is a technique that allows detection of G proteinmediated signaling in living cells by measuring cytoskeleton movements derived from receptor activation [27][28][29][30]. Selective agonists led to a significant signal although simultaneous treatment with agonists did not lead to summation of effects.…”

Section: Methamphetamine Blocks Cb 1 R Function In Hek-293t Cells Expmentioning

confidence: 99%

Methamphetamine Blocks Adenosine A2A Receptor Activation via Sigma 1 and Cannabinoid CB1 Receptors

Casanovas

Reyes‐Resina

Lillo

et al. 2021

IJMS

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Methamphetamine is, worldwide, one of the most consumed drugs of abuse. One important side effect is neurodegeneration leading to a decrease in life expectancy. The aim of this paper was to check whether the drug affects one of the receptors involved in neurodegeneration/neuroprotection events, namely the adenosine A2A receptor (A2AR). First, we noticed that methamphetamine does not affect A2A functionality if the receptor is expressed in a heterologous system. However, A2AR becomes sensitive to the drug upon complexes formation with the cannabinoid CB1 receptor (CB1R) and the sigma 1 receptor (σ1R). Signaling via both adenosine A2AR and cannabinoid CB1R was affected by methamphetamine in cells co-expressing the two receptors. In striatal primary cultures, the A2AR–CB1R heteromer complex was detected and methamphetamine not only altered its expression but completely blocked the A2AR- and the CB1R-mediated activation of the mitogen activated protein kinase (MAPK) pathway. In conclusion, methamphetamine, with the participation of σ1R, alters the expression and function of two interacting receptors, A2AR, which is a therapeutic target for neuroprotection, and CB1R, which is the most abundant G protein-coupled receptor (GPCR) in the brain.

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Label‐Free Dynamic Mass Redistribution and Bio‐Impedance Methods for Drug Discovery

Cited by 10 publications

References 56 publications

Allosteric targeting of the FFA2 receptor (GPR43) restores responsiveness of desensitized human neutrophils

Allosteric targeting of the FFA2 receptor (GPR43) restores responsiveness of desensitized human neutrophils

Parallelized label-free monitoring of cell adhesion on extracellular matrix proteins measured by single colour reflectometry

Methamphetamine Blocks Adenosine A2A Receptor Activation via Sigma 1 and Cannabinoid CB1 Receptors

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