Prevention of cellular stress and inflammatory mediators by interference with intracellular signaling. BV-2 cells were pretreated for 1 hour with 1 μM compound C, 1 μM BAPTA-AM or 5 μM U-73122, or for 6 hours with pertussis toxin (PTX) (100 ng/mL) prior to incubation with 100 μM MPEP for 24 hours. (A) Intracellular reactive oxygen species (ROS) levels were determined using dihydroethidium (DHE). (B) Mitochondrial mass was measured using MitoTracker®Red CMXRos. (C) Mitochondrial superoxide levels were detected using MitoSOX Red. The mRNA expression of inducible oxygen species (iNOS) (D) and IL-6 (E) were determined by quantitative PCR and values were normalized to GAPDH mRNA control. (F) IL-6 protein expression was measured by ELISA. Results represent the mean ± SD of at least three independent experiments. Significance was tested by one-way ANOVA followed by Tukey’s test. *P <0.05, **P <0.01; ***P <0.005. GAPDH, glyceraldehyde 3-phosphate dehydrogenase; MPEP, 2-methyl-6-(phenylethynyl)-pyridine.