SARS-CoV-2 N Protein Induces Acute Lung Injury in Mice via NF-ĸB Activation

Front Immunol. 2021 Dec 7:12:791753. doi: 10.3389/fimmu.2021.791753. eCollection 2021.

Abstract

Background: Infection of SARS-CoV-2 may cause acute respiratory syndrome. It has been reported that SARS-CoV-2 nucleocapsid protein (N-protein) presents early in body fluids during infection. The direct involvement of N-protein in lung injury is poorly understood.

Methods: Recombinant N-protein was pretreated with polymyxin B, a lipopolysaccharide (LPS)-neutralizing agent. C57BL/6, C3H/HeJ (resistant to LPS), and C3H/HeN (control for C3H/HeJ) mice were exposed to N-protein via intratracheal administration to examine acute lung injury. In vitro, bone marrow-derived macrophages (BMDMs) were cultured with N-protein to study phosphorylation of nuclear factor kappa B (NF-ĸB) p65, macrophage polarization, and expression of proinflammatory cytokines.

Results: N-protein produced acute lung injury in C57BL/6 mice, with elevated protein permeability, total cell count, neutrophil infiltration, and proinflammatory cytokines in the bronchioalveolar lavage. N-protein also induced lung injury in both C3H/HeJ and C3H/HeN mice, indicating that the effect could not be attributed to the LPS contamination. N-protein triggered phosphorylation of NF-ĸB p65 in vitro, which was abolished by both N-protein denaturation and treatment with an antibody for N-protein, demonstrating that the effect is N-protein specific. In addition, N-protein promoted M1 macrophage polarization and the expression of proinflammatory cytokines, which was also blocked by N-protein denaturation and antibody for N-protein. Furthermore, N-protein induced NF-ĸB p65 phosphorylation in the lung, while pyrrolidine dithiocarbamate, an NF-ĸB inhibitor, alleviated the effect of N-protein on acute lung injury.

Conclusions: SARS-CoV-2 N-protein itself is toxic and induces acute lung injury in mice. Both N-protein and NF-ĸB pathway may be therapeutic targets for treating multi-organ injuries in Coronavirus disease 2019 (COVID-19).

Keywords: COVID-19; NF- kappa B; SARS-CoV-2; acute lung injury; nucleocapsid (N) protein.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acute Lung Injury / metabolism
  • Acute Lung Injury / virology*
  • Animals
  • COVID-19*
  • Coronavirus Nucleocapsid Proteins / toxicity*
  • Mice
  • Mice, Inbred C3H
  • Mice, Inbred C57BL
  • NF-kappa B / metabolism*
  • Phosphoproteins / toxicity
  • SARS-CoV-2

Substances

  • Coronavirus Nucleocapsid Proteins
  • NF-kappa B
  • Phosphoproteins
  • nucleocapsid phosphoprotein, SARS-CoV-2