MiR-520b inhibits endothelial activation by targeting NF-κB p65-VCAM1 axis

Biochem Pharmacol. 2021 Jun:188:114540. doi: 10.1016/j.bcp.2021.114540. Epub 2021 Apr 2.

Abstract

MiR-520b belongs to the miR-373/520 family, is expressed only in human and nonhuman primates. Previous reports indicated that the expression of miR-520b was repressed in human atherosclerotic plaque tissue compared with healthy vessels. However, the role of miR-520b in coronary artery disease still remains to be uncovered. In this study, we demonstrated that endothelial cells (ECs) in human atherosclerotic plaques expressed miR-520b and aimed to elucidate the impact of miR-520b on EC activation and inflammatory response. To determine the potential targets of miR-520b, we performed RNA-seq analysis by transfecting miR-520b mimics in ECs. The quantitative real-time PCR (qPCR) validation suggested that miR-520b over-expression reduced pro-inflammatory gene expression (e.g. ICAM1, VCAM1, SELE) while the inhibition of miR-520b induced their expression. By combining bioinformatics prediction and functional assays, we identified that RELA (Nuclear Factor-κB (NF-κB) Transcription Factor P65) was a direct target of miR-520b. Moreover, miR-520b mimics attenuated monocyte adhesion and monocyte trans-endothelial migration (the initial steps of atherosclerotic formation) in response to lipopolysaccharides (LPS) stimulation. Re-expression of a non-miR-targetable version of p65 could rescue the reduced monocyte cell attachment, suggesting that this process is NF-κB p65 dependent. MiR-520b reduced the abundance of NF-κB p65 in cytoplasmic fractions without corresponding increase in nuclear fractions, indicating that this regulation is independent of p65 translocation process. MiR-520b mimics attenuated the activity of VCAM-1 promoter, whereas miR-520b inhibitor activated its activity. However, miR-520b inhibitor had no effect on promoter activity containing the mutated NF-κB p65 binding sites, strongly demonstrating that the impact of miR-520b on VCAM1 gene is mediated by NF-κB p65. Thus, we concluded that miR-520b suppressed EC inflammation and the cross-talk between monocytes and ECs by down-regulating NF-κB p65-ICAM1/VCAM1 axis and might serve as a potential therapeutic target for EC dysfunction and atherosclerosis.

Keywords: Cell adhesive molecules; Monocyte adhesion; Monocyte trans-endothelial migration; NF-κB p65; miR-520b.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Atherosclerosis / metabolism
  • Atherosclerosis / pathology
  • Base Sequence
  • Endothelial Cells / metabolism*
  • HeLa Cells
  • Human Umbilical Vein Endothelial Cells / metabolism
  • Humans
  • MicroRNAs / metabolism*
  • NF-kappa B / antagonists & inhibitors
  • NF-kappa B / metabolism*
  • THP-1 Cells
  • Transcription Factor RelA / antagonists & inhibitors
  • Transcription Factor RelA / metabolism*
  • Vascular Cell Adhesion Molecule-1 / antagonists & inhibitors
  • Vascular Cell Adhesion Molecule-1 / metabolism*

Substances

  • MIRN520 microRNA, human
  • MicroRNAs
  • NF-kappa B
  • Transcription Factor RelA
  • Vascular Cell Adhesion Molecule-1