Characterization of Primary Cilia Formation in Human ESC-Derived Retinal Organoids

Figure 2

Brn3b-positive RGCs lose primary cilia during later developmental stages of retinal development. (a) Confocal images of retinal organoids show RGCs visualized by Brn3b (red); primary cilia are identified by costaining for Arl13b (green) and Centrin3 (gray), markers for the ciliary axoneme and basal body, respectively. (b) Quantitative assessment confirms that ciliation at days 148 and 176 of retinal organoid development is significantly reduced compared to day 44. (c) Significant variations in ciliary length are not observed. (d) Calretinin-positive RGCs are readily identified by calretinin (green) coexpression with markers for RGCs (Brn3b, gray) and cilia (Arl13b, red). (e) At days 148 and 176 of retinal organoid development, ciliation is significantly decreased compared to day 44. (f) No significant changes in ciliary length are observed in calretinin-positive RGCs during retinal organoid development. Nuclei are stained with DAPI (blue). Scale bar: 10 μm; magnified, 1 μm. (g) Confocal images of day 148 retinal organoids show RGCs visualized by Brn3b (red), RGC subtype markers by Islet2 or Osteopontin (green), and primary cilia by Arl13b (gray).

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