State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, 555 Zuchongzhi Road, Shanghai 201203, China.
Steroids. 2013 Sep;78(9):813-22. doi: 10.1016/j.steroids.2013.04.018. Epub 2013 May 23.
15-Deoxy-Δ(12,14)-PGJ2 (15d-PGJ2) is one of the major metabolites from prostaglandin D2 in arachidonic acid (AA) metabolic pathway. It was determined as a ligand of peroxisome proliferator-activated receptor γ (PPARγ) functioning potently in adipocyte development. However, the fact that 15d-PGJ2 exerts also PPARγ-independent biological actions has highly addressed its multi-target behavior. Here, we identified that 15d-PGJ2 was an antagonist of farnesoid X receptor (FXR), as investigated by surface plasmon resonance, fluorescence quenching and homo time-resolved fluorescence based analyses, and the coactivator-recruitment and luciferase-reporter related investigation. Assay of 15d-PGJ2 regulation on hFXRα target genes revealed that treatment of HepG2 cells with 15d-PGJ2 resulted in the stimulation of mRNA expressions of bile-salt export pump (BSEP), and the decrease of cholesterol 7a-hydroxylase (CYP7a1). In addition, functional assays indicated that 15d-PGJ2 promoted the conversion of cholesterol to bile acids in HepG2 cells. Moreover, molecular docking combined with molecular dynamics simulation was applied to develop the possible model of 15d-PGJ2 binding to hFXRα ligand binding domain (LBD) at atomic level, and the responsible residues for 15d-PGJ2/hFXRα-LBD interaction were thereby determined, which were further confirmed by SPR assays against hFXRα-LBD site-directed mutations. Given that hFXRα functions potently in the regulation of hepatic bile acid metabolism and lipid/glucose homeostasis, our current work is expected to help better understand the multi-target features of this PGD2 metabolite in biological pathways, and 15d-PGJ2 as a new discovered FXR antagonist might find its potential application in further anti-hypercholesterol research.
15-脱氧-Δ(12,14)-PGJ2(15d-PGJ2) 是花生四烯酸 (AA) 代谢途径中前列腺素 D2 的主要代谢产物之一。它被确定为过氧化物酶体增殖物激活受体 γ (PPARγ) 的配体，在脂肪细胞发育中具有很强的作用。然而，15d-PGJ2 还发挥 PPARγ 非依赖性的生物学作用这一事实，高度表明了其多靶点行为。在这里，我们通过表面等离子体共振、荧光猝灭和同相时间分辨荧光分析以及共激活剂募集和荧光素酶报告基因相关研究，鉴定出 15d-PGJ2 是法尼醇 X 受体 (FXR) 的拮抗剂。对 15d-PGJ2 调节 hFXRα 靶基因的测定表明，用 15d-PGJ2 处理 HepG2 细胞导致胆汁盐输出泵 (BSEP) 的 mRNA 表达增加，胆固醇 7a-羟化酶 (CYP7a1) 减少。此外，功能测定表明 15d-PGJ2 促进了 HepG2 细胞中胆固醇向胆汁酸的转化。此外，应用分子对接结合分子动力学模拟，在原子水平上构建了 15d-PGJ2 与 hFXRα 配体结合域 (LBD) 结合的可能模型，并确定了负责 15d-PGJ2/hFXRα-LBD 相互作用的残基，这些残基进一步通过针对 hFXRα-LBD 定点突变的 SPR 测定得到证实。鉴于 hFXRα 在调节肝内胆汁酸代谢和脂质/葡萄糖稳态方面具有很强的作用，我们目前的工作有望帮助更好地理解这种 PGD2 代谢物在生物途径中的多靶点特征，而 15d-PGJ2 作为一种新发现的 FXR 拮抗剂，可能在进一步的抗高胆固醇研究中找到其潜在的应用。
