Differential signaling of cysteinyl leukotrienes and a novel cysteinyl leukotriene receptor 2 (CysLT) agonist, N-methyl-leukotriene C, in calcium reporter and beta arrestin assays.

摘要

The cysteinyl leukotrienes (cysLTs) LTC, LTD, and LTE are lipid mediators with physiological and pathophysiological functions. They exert their effects through G protein-coupled receptors (GPCRs), most notably via CysLT and CysLT receptor. The roles of the CysLT receptor are beginning to emerge. Both LTC and LTD are potent agonists for the CysLT receptor; however, LTC is rapidly converted to LTD, which is also the main endogenous ligand for the CysLT receptor. A selective and potent agonist at the CysLT receptor would facilitate studies to discern between receptor subtypes. We show here that N-methyl LTC (NMLTC), a metabolically stable LTC mimetic, is a potent and selective CysLT receptor agonist. Two expression systems were used to evaluate the functional activity of NMLTC at human and/or mouse CysLT and CysLT receptors. Through the aequorin cell-based assay for calcium-coupled GPCRs, NMLTC was almost equipotent to LTC at CysLT receptors but was the least efficacious at CysLT receptors. In a beta-galactosidase-beta-arrestin complementation assay, the human (h) CysLT receptor can couple with beta-arrestin-2, and NMLTC is slightly more potent for eliciting beta-arrestin-2 binding compared with cysLTs. Furthermore, LTE is nearly inactive in this assay compared with its weak partial agonist activity in the aequorin system. In a vascular leakage assay, NMLTC is potent and active in mice overexpressing hCysLT receptor in endothelium, whereas the response is abrogated in CysLT receptor knockout mice. Therefore, NMLTC is a potent subtype selective agonist for the CysLT receptor in vitro and in vivo, and it will be useful to elucidate its biological roles.