The Key Laboratory of Modern Toxicology, Ministry of Education, Center for Global Health, School of Public Health, Nanjing Medical University, Nanjing, China; The Key Laboratory of Reproductive Medicine, Institute of Toxicology, Nanjing Medical University, Nanjing, China.
Sir Run Run Hospital, Nanjing Medical University, Nanjing, China.
Environ Pollut. 2020 Sep;264:114723. doi: 10.1016/j.envpol.2020.114723. Epub 2020 May 3.
Di-n-butyl phthalate (DBP) is ubiquitously in the environment and has been detected in almost all of human bodies. Few data could be found about the effects of DBP on cardiovascular system, though its reproductive toxicities have been studied extensively. This study aimed to explore the effects of DBP on lipid metabolism, a key step during the formation of atherosclerosis, since DBP was recently reported to be associated with atherosclerosis. THP-1 macrophages were employed and exposed to various levels of DBP (10, 10, 10, 10 and 10 mol/L) or DMSO as control. Lipid accumulation was determined by detection of cellular total cholesterol, free cholesterol, cholesterol ester and content of lipid drops. Expressions of mRNA/miRNAs and proteins were measured by qRT-PCR and western blotting, respectively. Bioinformatic analysis and dual luciferase reporter assay were used to analyze the combination between miR200c-5p and ATP-binding cassette transporter A1 (ABCA1). Cholesterol efflux assay was executed to study the inhibitory effects of DBP on cholesterol efflux capability. Results revealed that DBP at 10 mol/L prompted THP-1 macrophages lipid accumulation by inhibiting cholesterol efflux via suppressing ABCA1 expression. In addition, a non-linear inverted U-shaped relationship between DBP and lipid accumulation could be observed. Moreover, miR200c-5p could directly targets to ABCA1 3'UTR and modulate ABCA1 expression. Besides, downregulation of ABCA1 expression and reduction of lipid efflux induced by DBP were due to the miR200c-5p upregulation. Collectively, these data suggested that DBP at levels relative to human exposure could increase lipid accumulation in THP-1 macrophages by decreasing cholesterol efflux through miR200c-5p-ABCA1, then potentiate the formation of atherosclerosis.
邻苯二甲酸二正丁酯（DBP）广泛存在于环境中，几乎存在于所有人的体内。尽管其生殖毒性已被广泛研究，但关于 DBP 对心血管系统影响的数据却很少。本研究旨在探讨 DBP 对脂代谢的影响，因为 DBP 最近被报道与动脉粥样硬化有关，而脂代谢是动脉粥样硬化形成的关键步骤。本研究采用 THP-1 巨噬细胞，并用不同浓度（10、10、10、10 和 10 mol/L）的 DBP 或 DMSO 作为对照进行处理。通过检测细胞总胆固醇、游离胆固醇、胆固醇酯和脂滴含量来确定脂类积累。通过 qRT-PCR 和 Western blot 分别测量 mRNA/miRNA 和蛋白质的表达。生物信息学分析和双荧光素酶报告基因检测用于分析 miR200c-5p 与三磷酸腺苷结合盒转运体 A1（ABCA1）之间的结合。胆固醇外排实验用于研究 DBP 对胆固醇外排能力的抑制作用。结果表明，DBP 在 10 mol/L 浓度下通过抑制 ABCA1 表达来抑制胆固醇外排，从而促进 THP-1 巨噬细胞的脂类积累。此外，还可以观察到 DBP 与脂类积累之间存在非线性倒 U 型关系。此外，miR200c-5p 可以直接靶向 ABCA1 3'UTR 并调节 ABCA1 表达。此外，DBP 下调 ABCA1 表达和减少脂质外排是由于 miR200c-5p 的上调。总之，这些数据表明，在相对人体暴露水平下，DBP 通过 miR200c-5p-ABCA1 减少胆固醇外排，从而增加 THP-1 巨噬细胞中的脂类积累，进而促进动脉粥样硬化的形成。
