Figure 4.

Ataluren increases the number of ciliated patient-derived fibroblasts. (A) Workflow. Healthy control human-derived fibroblasts and USH2A^G3142* patient-derived fibroblasts were seeded. 24 h later culture media was changed to starvation media. Cells were treated with dimethyl sulfoxide (DMSO), Gentamicin (1 mg/mL), or Ataluren (5 µg/µL), respectively. After 24 h cells were subjected for immunofluorescence analysis and the number of ciliated cells was counted. Four independent experiments were included. (B) Anti-acetylated tubulin (Actub, green) was used as a ciliary marker, Pericentrin 2 (PCTN2, red) as basal body marker and DAPI to stain the nucleus. All images are in the same magnification, scale bar represents: 25 µm. (C) The graph represents the percentage of ciliated cells versus the total number of cells in healthy control (light grey) and in patient-derived fibroblasts (dark grey), respectively. A significant decrease in the percentage of ciliated cells was observed in untreated, DMSO- or Gentamicin-(Gent)-treated USH2A^G3142* patient-derived cells versus healthy-donor derived fibroblasts. Ataluren treatment (5 µg/µl) restored the percentage of ciliated cells of patient-derived cells similar to that of healthy donors.