NBS1 is induced in macrophages after proinflammatory and M-CSF stimulation. Seven-day BMDMs were starved of growth factors for 16 to 18 hours and then incubated with different stimuli. IFN-γ (10 ng/mL) and LPS (10 ng/mL) were used as proinflammatory stimuli, IL-4 (10 ng/mL) as anti-inflammatory stimulus, and M-CSF (10 ng/mL) and GM-CSF (10 ng/mL) as growth factors. (A) Relative mRNA expression was determined at the indicated times after treatment. Each graph represents triplicate samples, and the results are shown as mean ± standard deviation (SD). (B) Protein levels of NBS1 were determined after 24 hours of stimulation. β-actin is included as a loading control. (C) Protein levels of other DNA damage response proteins were determined in macrophages upon stimulation with IFN-γ or M-CSF. β-actin is included as a loading control. (D) DNA damage was assessed by quantifying γH2AX foci. For each condition, 10 different fields with at least 14 cells each were quantified and the results are shown as number of foci per nucleus in each field. N-acetyl cysteine (NAC), a ROS scavenger, was used at 1 mM. All assays are representative of at least 3 independent experiments showing similar results. *P < .01 in relation to the controls when all the independent experiments had been compared.