miR-497-5p靶向沉默核苷酸结合寡聚化结构域样受体蛋白1促进巨噬细胞源性泡沫细胞胆固醇流出  

miR-497-5p promotes cholesterol efflux from macrophage-derived foam cells by targeting silencing nucleotide-binding oligomerization domain-like receptor protein 1

作　　者：陈孔 吴剑锋 杨宏发 刘阳 曾高峰 CHEN Kong;WU Jianfeng;YANG Hongfa;LIU Yang;ZENG Gaofeng(Department of Cardiology,the Second Affiliated Hospital of University of South China,Hengyang,Hunan 421001,China)

机构地区：[1]南华大学附属第二医院心血管内科,湖南省衡阳市421001

出　　处：《中国动脉硬化杂志》2020年第6期490-495,共6页Chinese Journal of Arteriosclerosis

基　　金：湖南省自然科学基金项目(2017JJ2224、2019JJ50532);湖南省卫生健康委科研计划课题(B2019108);湖南省教育厅课题(14C0988)。

摘　　要：目的观察miR-497-5p对核苷酸结合寡聚化结构域样受体蛋白1(NLRP1)靶向调控及其对细胞胆固醇流出的影响。方法生物信息学与荧光素酶报告基因验证miR-497-5p与NLRP1靶向结合。人源THP-1单核细胞经佛波酯及氧化低密度脂蛋白处理后成为泡沫细胞。使用miR-497-5p mimic和miR-497-5p inhibitor处理细胞。实时荧光定量PCR和Western blot检测NLRP1、含半胱氨酸的天冬氨酸蛋白酶1(Caspase-1)、凋亡相关斑点样蛋白(ASC)表达情况。酶联免疫吸附法测定细胞培养液白细胞介素1β(IL-1β)和IL-18含量。液体闪烁计数仪检测泡沫细胞胆固醇流出水平。高效液相色谱法检测泡沫细胞内脂质含量。结果 miR-497-5p mimic能够显著性降低野生型NLRP1 3′UTR报告基因的荧光素酶活性。与对照组相比,miR-497-5p mimic组NLRP1、ASC、Caspase-1的mRNA及蛋白表达水平明显下调,IL-1β和IL-18分泌明显减少。miR-497-5p mimic较对照组显著促进细胞胆固醇流出,减少细胞内总胆固醇、游离胆固醇和胆固醇酯的含量。结论 miR-497-5p可能通过靶向调控NLRP1,抑制巨噬细胞源性泡沫细胞炎症反应并促进胆固醇流出。Aim To observe the targeting regulation of mi R-497-5 p on nucleotide-binding oligomerization domain-like receptor protein 1( NLRP1) and its effect on cholesterol efflux. Methods Bioinformatics and luciferase reporter gene assay were used to verify the targeted binding of mi R-497-5 p and NLRP1. Human THP-1 monocytes were induced into foam cells after treatment with phorbol ester and oxidized low density lipoprotein. mi R-497-5 p mimic and mi R-497-5 p inhibitor were used to treat cells. Real-time fluorescence quantitative PCR and Western blot were used to detect the expressions of NLRP1,cysteinyl aspartate specific proteinase 1( Caspase-1) and apoptosis-associated speck-like protein containing a Caspase-recruitment domain( ASC). The contents of interleukin-1β( IL-1β) and IL-18 in cell culture medium were determined by enzyme-linked immunosorbent assay. Cholesterol efflux detection was performed by liquid scintillator. Lipid content in foam cells was detected by high performance liquid chromatography. Results The luciferase activity of wild type NLRP1 3’ UTR reporter gene was significantly reduced by mi R-497-5 p mimic. Compared with the control group,the m RNA and protein expression levels of NLRP1,ASC and Caspase-1 in mi R-497-5 p mimic group were significantly down regulated,and the secretions of IL-1β and IL-18 was significantly reduced. Compared with the control group,mi R-497-5 p mimic significantly promoted the cellular cholesterol efflux and reduced the contents of total cho-lesterol,free cholesterol and cholesterol ester in cells. Conclusion mi R-497-5 p may inhibit the inflammatory response of macrophage-derived foam cells and promote cholesterol efflux by targeting regulation of NLRP1.

关 键 词：miR-497-5p 核苷酸结合寡聚化结构域样受体蛋白1 胆固醇流出 脂质蓄积 动脉粥样硬化 

分 类 号：R54[医药卫生—心血管疾病] R363[医药卫生—内科学]