Figure 6.
Morphological analysis of alveolar capillaries. (A) Representative electron micrographs of intact and impaired endothelial junctions. The percentages of impaired endothelial junctions in the alveolar capillaries of control, LPS-treated, or LPS + Riociguat–treated mice were quantified. Scale bar: 0.5 μm. The data are presented as mean ± SD, n = 9 sections from three mice. Statistical significance was determined by the Kruskal–Wallis test. (B) Representative electron micrograph of caveolae in alveolar endothelium. The number of endothelial caveolae in the alveolar capillaries of control, LPS-treated, or LPS + Riociguat–treated mice were quantified. Scale bar: 0.5 μm. Data are presented as mean ± SD, n = 9 sections from three mice. Statistical significance was determined by the Kruskal–Wallis test. (C) Representative electron micrographs of intact and disrupted alveolar endothelium with small or large lesions. Pseudocolors highlight EC (red) and basement membrane (green). Arrowheads indicate the breakdown of the endothelium. Scale bar: 1 μm. Data are presented as mean ± SD, n = 9 sections from three mice. Statistics were performed using the Kruskal–Wallis test. (D) Representative electron micrographs showing the normal and detached pericyte cellular process over the endothelium. Pseudocolors highlight EC (red), basement membrane (green), and pericyte (blue). Arrowhead indicate a detached pericyte. Scale bar: 0.5 μm. Data are presented as mean ± SD, n = 9 sections from three mice. Statistics were performed using the Kruskal–Wallis test. (E) 3D reconstructed images of lung sections of Gucy1a1-EGFP::Tek-Cre::Rosa26-tdTomato mice treated with control, LPS, or LPS + Riociguat. Pericytes were labeled with EGFP, and ECs were labeled with tdTomato. Riociguat treatment rescues LPS-induced reduction of pericyte coverage. Scale bar: 5 μm. (F) Top panels are representative lung images of sparse-labeled Cspg4-CreERT2::Rosa26-tdTomato mice with PBS, LPS, or LPS + Riociguat treatment. Lower panels showing the 3D skeletonized pericytes with primary cytoplasmic processes and secondary cytoplasmic processes coded with red and blue, respectively. Scale bar: 20 μm. Violin plots showing the quantification of the mean length of primary and secondary cellular processes. Riociguat treatment suppressed LPS-induced cellular process retraction. Arrowheads indicate pericyte cytoplasmic processes. n = 20–30 sections from three mice. Statistical significance was determined by one-way ANOVA with Tukey test.

Morphological analysis of alveolar capillaries. (A) Representative electron micrographs of intact and impaired endothelial junctions. The percentages of impaired endothelial junctions in the alveolar capillaries of control, LPS-treated, or LPS + Riociguat–treated mice were quantified. Scale bar: 0.5 μm. The data are presented as mean ± SD, n = 9 sections from three mice. Statistical significance was determined by the Kruskal–Wallis test. (B) Representative electron micrograph of caveolae in alveolar endothelium. The number of endothelial caveolae in the alveolar capillaries of control, LPS-treated, or LPS + Riociguat–treated mice were quantified. Scale bar: 0.5 μm. Data are presented as mean ± SD, n = 9 sections from three mice. Statistical significance was determined by the Kruskal–Wallis test. (C) Representative electron micrographs of intact and disrupted alveolar endothelium with small or large lesions. Pseudocolors highlight EC (red) and basement membrane (green). Arrowheads indicate the breakdown of the endothelium. Scale bar: 1 μm. Data are presented as mean ± SD, n = 9 sections from three mice. Statistics were performed using the Kruskal–Wallis test. (D) Representative electron micrographs showing the normal and detached pericyte cellular process over the endothelium. Pseudocolors highlight EC (red), basement membrane (green), and pericyte (blue). Arrowhead indicate a detached pericyte. Scale bar: 0.5 μm. Data are presented as mean ± SD, n = 9 sections from three mice. Statistics were performed using the Kruskal–Wallis test. (E) 3D reconstructed images of lung sections of Gucy1a1-EGFP::Tek-Cre::Rosa26-tdTomato mice treated with control, LPS, or LPS + Riociguat. Pericytes were labeled with EGFP, and ECs were labeled with tdTomato. Riociguat treatment rescues LPS-induced reduction of pericyte coverage. Scale bar: 5 μm. (F) Top panels are representative lung images of sparse-labeled Cspg4-CreERT2::Rosa26-tdTomato mice with PBS, LPS, or LPS + Riociguat treatment. Lower panels showing the 3D skeletonized pericytes with primary cytoplasmic processes and secondary cytoplasmic processes coded with red and blue, respectively. Scale bar: 20 μm. Violin plots showing the quantification of the mean length of primary and secondary cellular processes. Riociguat treatment suppressed LPS-induced cellular process retraction. Arrowheads indicate pericyte cytoplasmic processes. n = 20–30 sections from three mice. Statistical significance was determined by one-way ANOVA with Tukey test.

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