Department of Cardiology, People's Hospital of Inner Mongolia Autonomous region, No. 20 of Zhaowuda Road, Hohhot city, 010020, Inner Mongolia, China.
The Center of Computer Information, People's Hospital of Inner Mongolia Autonomous region, Hohhot, 010020, Inner Mongolia, China.
Inflammation. 2020 Feb;43(1):66-73. doi: 10.1007/s10753-019-01094-y.
Our previous study has demonstrated that miR-23b enhances oxidized low-density lipoprotein (oxLDL)-induced inflammatory response of macrophages through the A20/NF-κB signaling pathway, thus contributing to atherosclerosis. This study aims to further investigate the upstream regulators of miR-23b in mediating oxLDL-induced inflammatory response. Human monocyte cell line THP1 was induced to differentiate into macrophages followed by the oxLDL stimulation of inflammatory response. The expression of miR-23b, LINC01140, and p53 mRNA was detected by quantitative PCR. The combination of miR-23b and LINC01140 was confirmed by luciferase reporter assay and RNA immunoprecipitation. The binding of p53 and LINC01140 promoter was determined by luciferase reporter assay. The level of inflammatory cytokines, including MCP-1, TNF-α, and IL-1β, was assessed by enzyme-linked immunosorbent assay. LINC01140 was downregulated, while p53 and miR-23b were upregulated in oxLDL-induced inflammatory response of macrophages. Overexpression of LINC01140 reduced NF-κB activity by reducing miR-23b and increasing A20. The transcription of LINC01140 was inhibited by binding of p53 and the LINC01140 promoter region. Knockdown of p53 significantly reduced NF-κB activity and level of inflammatory cytokines by promoting LINC01140 expression. Our findings demonstrated that LINC01140 acts as an anti-inflammatory factor through negatively regulating miR-23/A20 axis. In addition, p53 is identified as a transcriptional repressor of LINC01140.
我们之前的研究表明，miR-23b 通过 A20/NF-κB 信号通路增强氧化型低密度脂蛋白（oxLDL）诱导的巨噬细胞炎症反应，从而促进动脉粥样硬化。本研究旨在进一步探讨 miR-23b 在介导 oxLDL 诱导的炎症反应中的上游调控因子。用 oxLDL 刺激人单核细胞系 THP1 诱导分化为巨噬细胞，以诱导炎症反应。用实时定量 PCR 检测 miR-23b、LINC01140 和 p53 mRNA 的表达。用荧光素酶报告基因检测和 RNA 免疫沉淀实验验证 miR-23b 与 LINC01140 的结合。用荧光素酶报告基因检测实验确定 p53 与 LINC01140 启动子的结合。用酶联免疫吸附试验检测炎症细胞因子（包括 MCP-1、TNF-α 和 IL-1β）的水平。在 oxLDL 诱导的巨噬细胞炎症反应中，LINC01140 下调，而 p53 和 miR-23b 上调。LINC01140 的过表达通过降低 miR-23b 和增加 A20 来降低 NF-κB 活性。p53 通过与 LINC01140 启动子区域结合抑制 LINC01140 的转录。p53 敲低通过促进 LINC01140 表达显著降低 NF-κB 活性和炎症细胞因子水平。我们的研究结果表明，LINC01140 通过负调控 miR-23/A20 轴发挥抗炎作用。此外，p53 被鉴定为 LINC01140 的转录抑制因子。
