Figure 3

Figure 3

Saos-2 cells mimic CEP zebrafish model. (A, B) Mineralization in Saos-2 cells treated with MAC ± Uro-I was assayed using ARS staining (photograph, A; quantification, B). Staining was normalized to MAC only-treated cells (set to 100%). (C) Cell lysates from experiment in (A) were blotted with the indicated antibodies. (D) Quantification of LC3-II shown in (C). LC3-II level was normalized to MAC only (left panel) or vehicle-treated (right panel), set to 100%. (E) RT² Profiler PCR Array (left panel) and qPCR (right panel). Relative gene expression is represented as fold change normalized to housekeeping gene. Data are from 2 independent experiments. (F) Acitretin does not rescue reduced mineral matrix phenotype in uro-I-treated cells. ARS staining quantification as in (B). (G) Acitretin normalizes ER stress (BiP) and autophagy (LC3-II) markers. Dashed lines represent non-adjacent lanes in the gel. Coomassie-stained gel (C,G) shows equal protein loading. Full-length blots and gels (C,G) are presented in Fig.S4. (H) Quantification of LC3-II. LC3-II level was normalized to DMSO-treated cells set to 100%. (I) Gene expression profiling as in (E). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.