Li Yuanbin, Jiang Bimei, Liang Pengfei, Tong Zhongyi, Liu Meidong, Lv Qinglan, Liu Yanjuan, Liu Xuanyou, Tang Yuting, Xiao Xianzhong
Department of Pathophysiology, Xiangya School of Medicine, Central South University, Changsha, Hunan 410008, PR China.
Department of Pathophysiology, Xiangya School of Medicine, Central South University, Changsha, Hunan 410008, PR China.
Biochem Biophys Res Commun. 2017 Apr 29;486(2):364-371. doi: 10.1016/j.bbrc.2017.03.047. Epub 2017 Mar 14.
Our recent studies have indicated that nucleolin, as a multifunctional RNA-binding protein, exerts protective effects in the myocardial cells and endothelial cells under the condition of oxidative stress. However, the function of nucleolin and its potential mechanism in macrophage-derived foam cell formation remain largely unexplored. ApoE-/- mice were fed with a high-fat diet (HFD) for 10-24 weeks. Protein expression was measured by western blotting or immunofluorescence, and gene expression at the mRNA level was detected by qRT-PCR. The level of lipid in macrophages was examined by Oil Red O staining, high-performance liquid chromatography (HPLC) and NBD-cholesterol. Actinomycin D (Act D) was used to determine the stability of ABCA1 mRNA in macrophages. The interaction of nucleolin with ABCA1 mRNA was assessed using co-immunoprecipitation (co-IP). The aortas advanced plaques demonstrated significantly lower levels of nucleolin protein compared with early plaques in ApoE-/- mice, in which the macrophage foam cells occupied main body. Nucleolin expression at the mRNA and protein levels in RAW264.7 macrophages was significantly reduced by oxidized low-density lipoprotein (oxLDL) in a dose- and time-dependent manner. Furthermore, nucleolin overexpression markedly attenuated lipid accumulation in oxLDL-challenged macrophages through increasing cholesterol efflux. In addition, nucleolin overexpression significantly increased the expression of ATP-binding cassette transporter A1 (ABCA1) at the mRNA and protein levels without affecting expressions of scavenger receptors (SR)-A, SR-B1, CD36 and ATP-binding cassette transporter G1 (ABCG1) at the mRNA level. Moreover, nucleolin overexpression increased the stability of ABCA1 mRNA in macrophages, whereas nucleolin ablation abrogated the oxLDL-induced up-regulation of ABCA1. The up-regulation of ABCA1 by nucleolin resulted from its protein-RNA interaction. Our data suggested that nucleolin inhibited foam cell formation through enhancing stability of ABCA1 mRNA and subsequently increasing cholesterol efflux.
我们最近的研究表明，核仁素作为一种多功能RNA结合蛋白，在氧化应激条件下对心肌细胞和内皮细胞具有保护作用。然而，核仁素在巨噬细胞源性泡沫细胞形成中的功能及其潜在机制仍 largely 未被探索。给 ApoE - / - 小鼠喂食高脂饮食（HFD）10 - 24 周。通过蛋白质印迹或免疫荧光法测量蛋白质表达，通过 qRT - PCR 检测 mRNA 水平的基因表达。通过油红 O 染色、高效液相色谱（HPLC）和 NBD - 胆固醇检测巨噬细胞中的脂质水平。放线菌素 D（Act D）用于确定巨噬细胞中 ABCA1 mRNA 的稳定性。使用免疫共沉淀（co - IP）评估核仁素与 ABCA1 mRNA 的相互作用。在 ApoE - / - 小鼠中，主动脉晚期斑块中核仁素蛋白水平明显低于早期斑块，其中巨噬细胞泡沫细胞占主体。氧化低密度脂蛋白（oxLDL）以剂量和时间依赖性方式显著降低 RAW264.7 巨噬细胞中核仁素在 mRNA 和蛋白质水平的表达。此外，核仁素过表达通过增加胆固醇流出显著减轻 oxLDL 刺激的巨噬细胞中的脂质积累。此外，核仁素过表达在 mRNA 和蛋白质水平上显著增加 ATP 结合盒转运体 A1（ABCA1）的表达，而在 mRNA 水平上不影响清道夫受体（SR）-A、SR - B1、CD36 和 ATP 结合盒转运体 G1（ABCG1）的表达。此外，核仁素过表达增加了巨噬细胞中 ABCA1 mRNA 的稳定性，而核仁素缺失消除了 oxLDL 诱导的 ABCA1 上调。核仁素对 ABCA1 的上调是由于其蛋白质 - RNA 相互作用。我们的数据表明，核仁素通过增强 ABCA1 mRNA 的稳定性并随后增加胆固醇流出抑制泡沫细胞形成。
