Fig. 5.
Autophagy-deficient [hypomorphic (hm)] autophagy-related 16-like 1-deficient (hm) Atg16L1hm/hm mice have abnormal club cells. A–D: representative images of small airways from Atg16l1+/+and Atg16l1hm/hm littermate mice stained with hematoxylin and eosin (A and B) and periodic acid-Schiff (C and D). Scale bars = 100 µm. Insets show regions within black boxes (scale bars = 10 µm). Images of similar magnification from parallel sections stained with secretoglobulin family 1A member 1 (Scgb1a1) and acetylated α-tubulin (AAT) are shown below hematoxylin-and-eosin insets. Arrows indicate ciliated cells; arrowheads indicate club cells. E–H: localization of club cell proteins in airways of Atg16l1+/+ and Atg16l1hm/hm mice. Lung sections were immunostained as indicated for Scgb1a1 (green), surfactant protein A1 (Sftpa1, red), surfactant protein D (Sftpd, green), and cytochrome P-450 family 2, subfamily f, polypeptide 2 (Cyp2f2, red), and nuclei were stained with bis-benzamide (blue). Scale bars = 10 µm. I: percentage of high and sparse Scgb1a1-positive airway cells. Scgb1a1-positive cells were subdivided into those with high or sparse amounts of cytoplasmic puncta (see materials and methods). Values are means ± SE; n = 4–6 mice/group performed in 3 independent experiments; n = 4–6 well-orientated airways/mouse. **P < 0.01 (by one-way ANOVA with Tukey’s multiple comparison test). J: percentage of club cells that are positive for the indicated markers in Atg16l1+/+ and Atg16l1hm/hm mice. Values are means ± SE; n = 4–6 mice/group performed in 3 independent experiments; n = 4–6 well-orientated airways/mouse. **P < 0.01 (by Mann-Whitney U-test); ns, not significant.