Figure 5.

P. aeruginosa QS molecule induces ROS generation and oxidative mtDNA injury in bronchial epithelial cells. (A–C) BEAS-2B cells were treated with 3-oxo-C12-HSL (10 μM, 100 μM), or PAO1 (MOI 20), or a positive control (rotenone and antimycin A, R/A) for 6 hours. MitoSOX fluorescence was measured using flow cytometry. (A) Mean fluorescence intensity is displayed in a histogram format. Cells that stained positive for MitoSOX fluorescence (labeled Positive Cells) were quantified and compared between groups (B). 100 μM 3-oxo-C12-HSL resulted in a significant increase in ROS generation. (C,D) The ratio of a 79 bp mtDNA fragment to 230 bp mtDNA fragment was used to quantify mtDNA oxidative damage in BEAS-2B cells (C) and primary NhBEs (D) treated with 100 μM 3-oxo-C12-HSL or PAO1 for 6 hours. 100 μM 3-oxo-C12-HSL produced significant mtDNA oxidative injury in both BEAS-2B and NhBEs. A-C, Results are mean ± SEM. *p < 0.05, ***p < 0.001 by one-way ANOVA with Tukey’s multiple comparisons test. n = 3 independent experiments. (D) Results are mean ± SEM. *p < 0.05, unpaired t test, n = 3 independent experiments