Fig 2: Deletion of Ctnnb1 or Fgf10 in ACTA2+ Cells after Naphthalene Injury Impairs Epithelial Regeneration(A and F) Experimental setup and timeline of tamoxifen and NA treatment. Mice were fed tamoxifen-containing food.(B–D and G–I) Weight-loss curves (B and G), immunofluorescence (C, H), and quantification of immunoreactivity (D, I) are shown.(E and J) qPCR on lung homogenates.Ctrl, control mice; Euth, euthanasia; Exp, experimental mice; NA, naphthalene; Tam, Tamoxifen. Scale bars: 50 µm. (B and E) n = 3 per group, except n = 5 for Ctrl-Oil and n = 5 for Ctrl-NA d14; (C, D, H, and I) n = 3 per group; (G and J) n = 5 for Ctrl-Oil, n = 3 for Exp-Oil, n = 6 for Ctrl-NA d14, and n = 4 for Exp-NA d14. Data are represented as means ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. See also Figure S1.

Fig 3: Phenotypic analysis of E18.5 hearts with epicardial misexpression of human TBX2.(A) Histological analysis (n = 2) by hematoxylin and eosin staining of transverse sections of Tbx18cre/+;HprtCAG::TBX2/y hearts reveals pericardial defects (black arrow) but no further anomalies compared to the control (Tbx18cre/+). Scale bars are 500 µm. (B-D) Immunofluorescence analysis (n = 4) of indicated proteins on transverse sections of Tbx18cre/+ (control) and Tbx18cre/+;HprtCAG::TBX2/y hearts. Shown are magnified regions of the right ventricle. Scale bars are 100 µm. (B) Human TBX2 protein and TAGLN is found in the epicardium and in epicardium-derived cells in Tbx18cre/+;HprtCAG::TBX2/y hearts. Note that the level of human TBX2 expression is similar to the level of endogenous mouse TBX2 protein. (C) Coronary vessels are surrounded by ACTA2- and TAGLN-expressing SMCs in Tbx18cre/+;HprtCAG::TBX2/y hearts at E18.5, but additional ACTA2- and TAGLN-positive cells are detected in the myocardium of these hearts. (D) Small EMCN-positive coronary vessels form in comparable densities in Tbx18cre/+;HprtCAG::TBX2/y and control hearts. Intramyocardial deposition of POSTN was detected in both genotypes in a comparable fashion. CAr, coronary artery; Epi, epicardium; LV, left ventricle; Peri, pericardium; RA, right atrium; RV, right ventricle.

Fig 4: Loss of NEDD4-2 results in kidney injury in juvenile mice. (a) Representative images showing hematoxylin–eosin (H&E) staining. Pathological observations include enlarged tubules (*), cellular debris within tubules (arrows) and mesenchyme infiltration (arrowheads). (b) Immunostaining shows increased expression of vimentin (mesenchymal marker), (SMA; myofibroblasts) and KIM-1 (marker of proximal tubule kidney injury). Picrosirius red staining demonstrates fibrosis in Nedd4-2-/- kidneys. (c) Increased gene expression for markers of kidney pathology, including collagen (encoded by Col1a1), vimentin (Vim), SMA (Acta2) and Kim-1, as determined by qRT-PCR in Nedd4-2+/+ or Nedd4-2-/- mice. Data are normalized to tbp levels and represent mean±S.E.M. for four mice per genotype. *P<0.05. Scale bars: (a)=50 µm, inset=1 mm; (b) (vimentin, picrosirius red and SMA)=100 µm, (KIM-1)=50 µm

Fig 5: Progressive kidney injury in kidney-specific Nedd4-2 KO mice. (a) Representative hematoxylin–eosin (H&E) images show enlarged tubules (black asterisk), thinning of lining epithelium (open arrowhead), cellular debris within tubules (black arrows) and mesenchyme infiltration (black arrowhead) in P40 Nedd4-2Ksp1.3 mice. (b) Picrosirius red staining demonstrates fibrosis in Nedd4-2Ksp1.3kidneys (red). (c) Immunostaining shows increased expression of vimentin, SMA and Kim-1. (d) Increased gene expression for markers of kidney injury at P40, including collagen (encoded by Col1a1), vimentin (Vim), SMA (Acta2) and Kim-1, determined by qRT-PCR. Only Kim-1 levels were significantly altered at P20. Data are normalized to tbp levels and represent mean±s.e.m. for four mice per genotype. (e) Immune cell infiltration demonstrated by staining for CD3 (T-lymphocyte marker; and quantitated in (f)). (g and h) Immunostaining and quantitation of cleaved caspase-3-expressing cells shows apoptotic cells within tubules (white asterisk) and lining epithelia (white arrows) of Nedd4-2Ksp1.3kidneys at P40. (f and h) n=4 per genotype. *P<0.05, **P<0.01, ***P<0.005 and ****P<0.001. Scale bars: (a)=50 µm, inset=1 mm; (b)=100 µm; (c) vimentin and SMA=100 µm, Kim-1=50 µm; (e and g)=50 µm. fov, Field of view; NS=not significant (P>0.05)