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Establishment of a Cell Line Stably Expressing the Growth Hormone Secretagogue Receptor to Identify Crocin As a Ghrelin Agonist

Overview
Journal Biomolecules
Publisher MDPI
Date 2022 Dec 23
PMID 36551241
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Abstract

The growth hormone secretagogue receptor-1a (GHSR1a) is the endogenous receptor for ghrelin. Activation of GHSR1a participates in many physiological processes including energy homeostasis and eating behavior. Due to its transitory half-life, the efficacy of ghrelin treatment in patients is restricted; hence the development of new adjuvant therapy is an urgent need. This study aimed to establish a cell line stably expressing GHSR1a, which could be employed to screen potential ghrelin agonists from natural compounds. First, by means of lentiviral transduction, the genome of a human HEK293T cell was modified, and a cell platform stably overexpressing GHSR1a was successfully established. In this platform, GHSR1a was expressed as a fusion protein tagged with mCherry, which allowed the monitoring of the dynamic cellular distribution of GHSR1a by fluorescent microscopy. Subsequently, the authenticity of the GHSR1a mediated signaling was further characterized by using ghrelin and teaghrelin, two molecules known to stimulate GHSR1a. The results indicated that both ghrelin and teaghrelin readily activated GHSR1a mediated signaling pathways, presumably via increasing phosphorylation levels of ERK. The specific GHSR1a signaling was further validated by using SP-analog, an antagonist of GHSR1a as well as using a cell model with the knockdown expression of GHSR1a. Molecular modeling predicted that crocin might be a potential ghrelin agonist, and this prediction was further confirmed by the established platform.

References
1.
Sadakane C, Muto S, Nakagawa K, Ohnishi S, Saegusa Y, Nahata M . 10-Gingerol, a component of rikkunshito, improves cisplatin-induced anorexia by inhibiting acylated ghrelin degradation. Biochem Biophys Res Commun. 2011; 412(3):506-11. DOI: 10.1016/j.bbrc.2011.08.002. View

2.
Jang Y, Kim T, Shim W . Naringin exhibits in vivo prokinetic activity via activation of ghrelin receptor in gastrointestinal motility dysfunction rats. Pharmacology. 2013; 92(3-4):191-7. DOI: 10.1159/000354579. View

3.
MKadmi C, Leyris J, Onfroy L, Gales C, Sauliere A, Gagne D . Agonism, Antagonism, and Inverse Agonism Bias at the Ghrelin Receptor Signaling. J Biol Chem. 2015; 290(45):27021-27039. PMC: 4646384. DOI: 10.1074/jbc.M115.659250. View

4.
Heppner K, Chaudhary N, Muller T, Kirchner H, Habegger K, Ottaway N . Acylation type determines ghrelin's effects on energy homeostasis in rodents. Endocrinology. 2012; 153(10):4687-95. PMC: 3512022. DOI: 10.1210/en.2012-1194. View

5.
Ramirez V, van Oeffelen W, Torres-Fuentes C, Chruscicka B, Druelle C, Golubeva A . Differential functional selectivity and downstream signaling bias of ghrelin receptor antagonists and inverse agonists. FASEB J. 2018; 33(1):518-531. DOI: 10.1096/fj.201800655R. View