Figure 5
From: TGFβ2-induced senescence during early inner ear development
IGF-1 modulates developmental senescence of chicken inner ear. (A–F) SAβG staining in isolated otic vesicles from stage HH18 embryos cultured ex vivo for 20 h in free serum medium (0S), in the presence of IGF-1 (10 nM), Sor (2.5 µM), AKTi (50 μM) or a combination. (G) Phosphorylation state of IGF-1 targets in HH18 otic vesicles explanted and incubated with the indicated treatments. pERK, pAKT and AKT were measured by western blotting of at least n = 15 otic vesicle lysates in two independent experiments. To improve the clarity of the presentation lanes have been grouped. Full-length blots are presented in Supplementary Figure S3. (H) SAβG staining quantification by FIJI software. Data are shown as mean SAβG area ± SEM with respect to the otic vesicle area, normalized to the 0S condition. *p < 0.05, ***p < 0.001 vs 0S; #p < 0.05, ###p < 0.001 vs. indicated inhibitor. (I) Igf1 and Igf1r mRNA levels from explanted HH18 otic vesicles in the absence (0S, white bars) or presence of palbociclib (1 µM). Gene expression was calculated as 2−ΔΔCt and normalized to the levels of the control condition (0S). The results are expressed as the means ± SEM from three independent experiments performed in triplicate. Statistical significance was determined with the Student’s t-test: ***P < 0.005, versus 0S. Ed, endolymphatic duct. Orientation: A, anterior; D, dorsal. Scale bar: 150 μm.
