Label-free dynamic mass redistribution analysis of endogenous adrenergic receptor signaling in primary preadipocytes and differentiated adipocytes

TY - JOUR

T1 - Label-free dynamic mass redistribution analysis of endogenous adrenergic receptor signaling in primary preadipocytes and differentiated adipocytes

AU - Jørgensen, Christinna V

AU - Zhou, Hong

AU - Seibel, Markus J

AU - Bräuner-Osborne, Hans

N1 - Crown Copyright © 2019. Published by Elsevier Inc. All rights reserved.

PY - 2019/5/1

Y1 - 2019/5/1

N2 - INTRODUCTION: Adipose tissues release adipokines, which regulate energy intake and expenditure. G protein-coupled receptors (GPCRs) and associated signaling pathways in adipocytes are potentially important drug targets for conditions with disturbed energy metabolism.METHODS: The aim of the current study was to compare signaling of endogenously expressed GPCRs between primary preadipocytes and differentiated adipocytes using a novel state-of-the-art unbiased method that measures dynamic mass redistribution (DMR) in real-time. Adrenergic agonists were chosen since they control adipocyte functions such as lipolysis and glycogenolysis.RESULTS: Isoprenaline (ISO) and phenylephrine (PE) elicited concentration-dependent responses in preadipocytes and differentiated adipocytes. The effect of ISO was cholera toxin (CTX)-sensitive, indicating it is Gs-dependent. The effect could also be blocked by propranolol proving the signal is mediated through β-adrenergic receptors. The signaling resulting from PE stimulation was completely abolished by the Gq/11-selective inhibitor FR900359 and CTX in preadipocytes but surprisingly became FR900359-insensitive but remained CTX-sensitive in differentiated adipocytes. The use of prazosin and propranolol revealed that the PE-response in differentiated adipocytes had a β-adrenergic receptor component to it. In addition, we tested the bone-derived peptide osteocalcin, which did not result in DMR changes in preadipocytes or differentiated adipocytes.DISCUSSION: In conclusion, this study for the first time demonstrates that DMR assays can be used to assess signaling in differentiated adipocytes. This platform can serve as a tool for future drug screening in primary adipocytes. Furthermore, this study illustrates that PE-induced effects on adipocytes vary by developmental stage and are not as selective as originally thought.

AB - INTRODUCTION: Adipose tissues release adipokines, which regulate energy intake and expenditure. G protein-coupled receptors (GPCRs) and associated signaling pathways in adipocytes are potentially important drug targets for conditions with disturbed energy metabolism.METHODS: The aim of the current study was to compare signaling of endogenously expressed GPCRs between primary preadipocytes and differentiated adipocytes using a novel state-of-the-art unbiased method that measures dynamic mass redistribution (DMR) in real-time. Adrenergic agonists were chosen since they control adipocyte functions such as lipolysis and glycogenolysis.RESULTS: Isoprenaline (ISO) and phenylephrine (PE) elicited concentration-dependent responses in preadipocytes and differentiated adipocytes. The effect of ISO was cholera toxin (CTX)-sensitive, indicating it is Gs-dependent. The effect could also be blocked by propranolol proving the signal is mediated through β-adrenergic receptors. The signaling resulting from PE stimulation was completely abolished by the Gq/11-selective inhibitor FR900359 and CTX in preadipocytes but surprisingly became FR900359-insensitive but remained CTX-sensitive in differentiated adipocytes. The use of prazosin and propranolol revealed that the PE-response in differentiated adipocytes had a β-adrenergic receptor component to it. In addition, we tested the bone-derived peptide osteocalcin, which did not result in DMR changes in preadipocytes or differentiated adipocytes.DISCUSSION: In conclusion, this study for the first time demonstrates that DMR assays can be used to assess signaling in differentiated adipocytes. This platform can serve as a tool for future drug screening in primary adipocytes. Furthermore, this study illustrates that PE-induced effects on adipocytes vary by developmental stage and are not as selective as originally thought.

U2 - 10.1016/j.vascn.2019.03.005

DO - 10.1016/j.vascn.2019.03.005

M3 - Journal article

C2 - 30946893

SN - 1056-8719

VL - 97

SP - 59

EP - 66

JO - Journal of Pharmacological and Toxicological Methods

JF - Journal of Pharmacological and Toxicological Methods

ER -