Hello everybody, I'm planning to investigate the in-vitro release of different markers (CXCL10, INF-a, IFN-b, 1L-1a, IL-1b, IL-6, free DNA) by NR8383 rat macrophages, upon incubation with several particles, in particular carbon nanotubes and silica.
Therefore, my question is: before expose cells to particles, do I have to activate/stimulate such macrophages with LPS, or is it an unnecessary step? Thank you.
Hey Riccardo,
I am only experienced with mouse macrophages, but macrophages will secrete Cytokines and Chemokines (also those you listed) just after sole treatment with LPS.
LPS as a bacterial stimulus will lead to cytokine production and secretion via TLR4 signaling. Normally, ex vivo macrophages do not need to be pushed into a pro-inflammatory state.
If you pre-treat your macrophages with LPS you will get for sure cytokines and chemokines, but you cannot tell, if your particles have this effect.
I would suggest, you treat your macrophages with your particles (one to three different concentrations) and one set with LPS, but just as a positive control. As time points you can take supernatants after 5 and 24 hours after treatment and analyse with ELISA or Procartaplex.
All the best and stay healthy,
Marc
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