摘要
目的观察糖基化终末产物(advanced glycation end products,AGEs)及其受体(receptor of advanced glycation end products,RAGE)对巨噬细胞胆固醇流出功能的影响。方法培养THP-1细胞株并用佛波酯诱导分化使其成为巨噬细胞,与100μg/ml的氧化LDL(oxidized LDL,ox LDL)孵育细胞使其转化为泡沫细胞。分别以浓度为300μg/ml的AGEs、600μg/ml的AGEs对细胞进行刺激,用浓度为10μg/ml的抗RAGE抗体对细胞进行预处理,通过油红"O"染色测定来反映细胞内脂质含量,通过RT-PCR及Western Blot的方法来检测各组巨噬细胞RAGE表达以及与胆固醇流出相关因子ATP-结合盒转运子(ATP-binding cassette,ABC)G1、ABCA1及肝X受体-α(Liver X Receptor-α,LXRα的表达变化),通过将干预后的巨噬细胞与荧光标记的ox LDL共同孵育,并加入HDL及载脂蛋白A1(apolipoprotein A1,apo A1)介导胆固醇流出,测定细胞培养基中的荧光强度,动态观察巨噬细胞在不同干预条件下胆固醇流出功能变化。结果经AGEs诱导后,巨噬细胞内脂质含量增加,RAGE表达增加,ABCG1表达下降,巨噬细胞胆固醇流出能力减弱。AGEs效应呈浓度依赖性。应用抗体阻断RAGE功能后,AGEs引起的改变均有明显恢复。结论 AGEs-RAGE相互作用可以抑制巨噬细胞流出胆固醇,增加细胞内脂质累积,从而易于形成泡沫细胞。
Objective To investigate the interaction of advanced glycation end products (AGEs) and receptor of advanced glycation end products (RAGE) in cholesterol efflux of macrophage. Methods Human THP-1 monocytes were treated with PMA( 100 ng/ml) for 48 h, and then induced with oxLDL( 100μg/ml) Dil-oxLDL(40 μg/ml) for differentiating to macrophage foam cells. Cells was pretreated by AGEs (300 μg/ml or 600 μg/ml) for 2 h, and pre-stimulated with antibody for RAGE (10 μg/ml). The staining of oil red O and testing of cholesterol ester in macrophages were used for evaluating the accumulation of lipid in macrophages. RT-PCR and Western blotting analysis were using to test the expression of RAGE, ABCG1, ABCA1 and LXR-α. After adding HDL and apoAI to mediate cholesterol efflux, the fluorescence intensity in the cell medium was tested to reflect the ability of cholesterol efflux in macrophages. Results AGEs can increase the content of lipid and RAGE expression in macrophages in a concentration-dependent manner. The ability of macrophage for cholesterol efflux was decreased accompanied with the reduction of ABCG1 expression after treating with AGEs. For blocking up the AGEs-RAGE axis by antibody for RAGE, all the changes were reversed. Conclusion AGEs-RAGE interaction can decrease cholesterol efflux in macrophages, which can easily transform to foam cell.
出处
《同济大学学报(医学版)》
CAS
2016年第6期23-28,63,共7页
Journal of Tongji University(Medical Science)
基金
国家自然科学基金青年项目(81300699)
上海市卫生与计划生育委员会青年科研项目(20124Y106)
上海市浦东新区优秀青年医学人才计划(PWRq2013-04)