Fig. 4

Effects of leflunomide (10µM) and BGP15 (15µM) on cell proliferation, oxidative stress, senescence-associated secretory phenotype (SASP) components mRNA levels and mitochondrial morphology in A549 cells in the presence and absence of cigarette smoke extract (CSE). Cell viability (A), cell proliferation (B), intracellular ROS (C) and mitochondrial ROS (D) are respectively detected by CCK-8, Edu staining, DCFH-DA and Mito SOX Red, and data are presented as percentage changes compared to control cells. Quantitative real-time PCR is employed to measure the mRNA levels of IL-1β (E), IL-6 (F), CXCL1 (G) and CXCL8 (H) in relation to β-actin, and data are presented as individual and mean values of fold changes compared to control cells. N = 8 in each group. Representative confocal microscopy images of the mitophagy dye fluorescence and Mito-Tracker Green staining (I, original magnification, ×63). The scale bar is 10 μm. The fluorescent intensity of mitophagy (J), the area of mitochondria (K), and percentage of mitochondrial fragmentation (L) and perinuclear mitochondrial compaction (M) are assessed by MitoTracker Green. Representative fluorescent microscopy images of JC-1 staining (N) and JC-1 values (O). The scale bar is 50 μm. N = 8 in each group. Data are presented as individual and mean values. ^*p < 0.05, ^**p < 0.01, ^***p < 0.001