Induction of inducible nitric oxide synthase (iNOS) in Porphyromonas gingivalis LPS-treated mouse macrophage cell line (RAW264.7) requires Toll-like receptor 9

Inflamm Res. 2018 Sep;67(9):723-726. doi: 10.1007/s00011-018-1168-1. Epub 2018 Jul 6.


Objective: The aim of this study is to investigate the involvement of TLR9 in the regulation of iNOS expression and nitric oxide (NO) production in Porphyromonas gingivalis LPS-treated mouse macrophages.

Methods: Mouse macrophage cell line (RAW264.7) was transfected with siRNAs against TLR9 and then stimulated with P. gingivalis LPS. At indicated time points, the activated cells were lysed. Gene and protein expression of iNOS were determined by RT-PCR and immunoblotting, respectively. The level of nitric oxide (NO) production in the supernatant of the activated cells was determined by Griess reaction assay.

Results and conclusion: Depletion of TLR9 in mouse macrophages demonstrated the markedly decreased iNOS gene and protein expression by P. gingivalis LPS compared to those of the wild-type or control siRNA transfected cells. In consistent with these results, the level of NO secretion was also significantly diminished in TLR9-depleted cells after challenged with P. gingivalis LPS. These results indicate that TLR9 involves in the regulation of the iNOS expression and the NO secretion in P. gingivalis LPS-treated macrophages.

Keywords: Nitric oxide; Porphyromonas gingivalis LPS; RAW264.7; TLR9; iNOS.

MeSH terms

  • Animals
  • Enzyme Induction / drug effects
  • Lipopolysaccharides / pharmacology*
  • Macrophages / drug effects*
  • Macrophages / metabolism
  • Mice
  • Nitric Oxide / metabolism*
  • Nitric Oxide Synthase Type II / genetics
  • Nitric Oxide Synthase Type II / metabolism*
  • Porphyromonas gingivalis
  • RAW 264.7 Cells
  • RNA, Messenger / metabolism
  • Toll-Like Receptor 9 / metabolism*


  • Lipopolysaccharides
  • RNA, Messenger
  • Tlr9 protein, mouse
  • Toll-Like Receptor 9
  • Nitric Oxide
  • Nitric Oxide Synthase Type II
  • Nos2 protein, mouse