Fig 2: Subchronic cigarette smoke exposure affects circadian focused mRNA expression analyzed by NanoString.Total RNA was isolated from the lungs of mice exposed to air and cigarette smoke (CS) for 30 days. Our customized NanoString panel (circadian gene focused) was used to screen the potential targets via nCounter SPRINT Profiler. Normalization of absolute RNA count and data analysis were done by nSolver software. The overview of all the dysregulated targets is shown as a heatmap (A), and selected gene transcription changes are shown separately (B). Data are shown as mean ± SEM (n = 6; *P < 0.05, **P < 0.01, ***P < 0.001 between groups; #P < 0.05, ##P < 0.01 compared with CS-exposed WT group; &&P < 0.01 compared with WT air controls, 1-way ANOVA with Šidák correction).

Fig 3: Transcript levels of senescence and mitochondrial function-associated genes are altered in PBS/GCV treated CS-exposed p16-3MR mice. Total RNA was isolated from the lungs of PBS/GCV treated and air/CS-exposed (3 months) p16-3MR mice. Our customized NanoString panel (senescence) was used to screen the potential targets via nCounter SPRINT Profiler. Changes in the normalized counts of respective genes were obtained relative to PBS-Air group and plotted as heat map. Heat map showing fold changes (FC) in the expression of (a) senescence and (b) mitochondrial function associated genes is graphed relative to PBS-Air group.

Fig 4: GCV treatment reverses the genes associated with EMT in CS-exposed p16-3MR mice. Total RNA was isolated from the lungs of PBS/GCV treated and air/CS-exposed (3 months) p16-3MR mice. Our customized NanoString panel to study lung regeneration was used to screen the potential targets via nCounter SPRINT Profiler. Normalization of absolute RNA count and data analysis were done by nSolver software. The overview of all the dysregulated targets is shown as a (a) heatmap and (b) selected gene transcription changes between PBS-CS versus GCV-CS are shown separately. Data are shown as mean ± SEM (n = 3/group); *p < 0.05, **p < 0.01 versus PBS-CS, per unpaired t test for pairwise comparison. Here; ns: not significant. (c) Schematics demonstrating how removal of p16+ senescent cells in CS (3 months)-exposed p16-3MR mice causes reduction in neutrophilic inflammation, CS-induced mitochondrial dysfunction and tissue damage.