Inhibiting nucleolin reduces inflammation induced by mitochondrial DNA in cardiomyocytes exposed to hypoxia and reoxygenation

Br J Pharmacol. 2019 Nov;176(22):4360-4372. doi: 10.1111/bph.14830. Epub 2019 Oct 27.


Background and purpose: Cellular debris causes sterile inflammation after myocardial infarction. Mitochondria constitute about 30 percent of the human heart. Mitochondrial DNA (mtDNA) is a damage-associated-molecular-pattern that induce injurious sterile inflammation. Little is known about mtDNA's inflammatory signalling pathways in cardiomyocytes and how mtDNA is internalized to associate with its putative receptor, toll-like receptor 9 (TLR9).

Experimental approach: We hypothesized that mtDNA can be internalized in cardiomyocytes and induce an inflammatory response. Adult mouse cardiomyocytes were exposed to hypoxia-reoxygenation and extracellular DNA. Microscale thermophoresis was used to demonstrate binding between nucleolin and DNA.

Key results: Expression of the pro-inflammatory cytokines IL-1β and TNFα were upregulated by mtDNA, but not by nuclear DNA (nDNA), in cardiomyocytes exposed to hypoxia-reoxygenation. Blocking the RNA/DNA binding protein nucleolin with midkine reduced expression of IL-1β/TNFα and the nucleolin inhibitor AS1411 reduced interleukin-6 release in adult mouse cardiomyocytes. mtDNA bound 10-fold stronger than nDNA to nucleolin. In HEK293-NF-κB reporter cells, mtDNA induced NF-κB activity in normoxia, while CpG-DNA and hypoxia-reoxygenation, synergistically induced TLR9-dependent NF-κB activity. Protein expression of nucleolin was found in the plasma membrane of cardiomyocytes and inhibition of nucleolin with midkine inhibited cellular uptake of CpG-DNA. Inhibition of endocytosis did not reduce CpG-DNA uptake in cardiomyocytes.

Conclusion and implications: mtDNA, but not nDNA, induce an inflammatory response in mouse cardiomyocytes during hypoxia-reoxygenation. In cardiomyocytes, nucleolin is expressed on the membrane and blocking nucleolin reduce inflammation. Nucleolin might be a therapeutic target to prevent uptake of immunogenic DNA and reduce inflammation.

Linked articles: This article is part of a themed section on Mitochondrial Pharmacology: Featured Mechanisms and Approaches for Therapy Translation. To view the other articles in this section visit

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • CpG Islands
  • DNA / metabolism*
  • Fibroblasts / metabolism
  • HEK293 Cells
  • Humans
  • Hypoxia / metabolism*
  • Inflammation / genetics
  • Inflammation / metabolism
  • Male
  • Mice, Inbred C57BL
  • Myocytes, Cardiac / metabolism*
  • NF-kappa B / metabolism
  • Oxygen / pharmacology
  • Phosphoproteins / genetics
  • Phosphoproteins / metabolism*
  • RNA-Binding Proteins / genetics
  • RNA-Binding Proteins / metabolism*
  • Toll-Like Receptor 9 / genetics
  • Toll-Like Receptor 9 / metabolism


  • NF-kappa B
  • Phosphoproteins
  • RNA-Binding Proteins
  • Toll-Like Receptor 9
  • nucleolin
  • DNA
  • Oxygen