Fig. 3

3D spheroid culture improved the viability of hiPSC-RPE and helped in maintaining RPE properties and functions. a A rubber micro-mold with multiple projections. b Agarose mold with imprint microwells. c Formation of hiPSC-RPE cell spheroid in agarose 3D Petri dishes. d Viability staining of dispersed hiPSC-RPE cell spheroid. Non-spheroid cultured hiPSCs were used as control. e Bar chart represents the percentage of dead cells of the control compared to 3D spheroid cultured hiPSC. f Immunoblotting analysis: the protein expression of ZO-1, RPE65, Nestin, and α-SMA in hiPSC-RPE after spheroid culturing. g Bar chart representing relative protein expression of several RPE-specific markers of 3D spheroid cultured hiPSC-RPE. h Schematic of TEER value measurements with the insert culture. i TEER assay results reveal that 3D spheroid culture could improve the hiPSC-RPE cells barrier function. Scale bar 200 μm (d). Data are presented as mean ± SEM. p values were determined by unpaired two-tailed Student’s t test (f), n = 5 for each group, *p < 0.05, **p < 0.01