Figure 4
Brillouin imaging of murine bladder walls in a model of actinic cystitis (X-ray radiation) at different times post-irradiation. (A) Schematic representation of the experiment: 2 months old rats were exposed to X-rays and then sacrificed at different timepoints. Each timepoint matched the healthy counterpart reported in Fig. 3. (B) Representative fibrotic bladder DIC image and Brillouin shift map of the whole bladder wall (urothelium – lamina propria – muscle) from tissues belonging to rats sacrificed 2, 4, or 6 months after the irradiation. Scale bars = 20 µm. Different layer boundaries are indicated by dotted black lines. (C) Kinetics of Brillouin shift distributions from X-ray irradiated bladders at different time points (green) and comparison to healthy animals of the same age (grey). N = 1–3 rats per time point and condition, each tissue layer was characterized by over 4 Brillouin maps of at least 150 × 40 μm2. (D) Brillouin shifts medians of irradiated rats (green) compared to control rats (grey). When compared with their age-matched controls (here shown as * marks), the Brillouin shift of month 2 U was significantly higher than its control (p = 0.0095), while at month 6 it was lower (p = 0.045); L was lower both at month 4 (p = 0.007) and 6 (p = 0.0003) after X-rays. When comparing the effects of radiation on different stages of fibrosis (here shown as ° marks), U showed a significant (p = 0.011) decrease from month 2 to 6, while L both from month 2 to 6 (p = 0.0014) and from month 4 to 6 (p = 0.018). No significant changes in the muscle layer have been observed. (Mean ± SEM. For comparing fibrotic with their age-matched healthy counterpart, we used Mann–Whitney U-test: *, **, ***: p < 0.05, p < 0.01, p < 0.001. For comparing the stages of fibrosis between different timepoints, we used Kruskall-Wallis test followed by Dunn’s multiple-comparison test: °, °°: p < 0.05, p < 0.01.).
