Fig. 5

Reproduction of Parkinson’s disease phenotypes using neurons derived from PARK2-LiPSCs. a An overview of the culture protocol used in this experiment. b Immunostaining of LiPSC (LPB10)-derived dopaminergic neurons with antibodies raised against the indicated markers. Scale bars = 100 μm. c Analysis of dopaminergic neuron differentiation by quantifying βIII-TUBULIN and TH double-positive neurons at Day 55 (n = 3 independent experiments; means ± SEM; **p < 0.01; Student’s t-test). d The deletion of exons 6 and 7 was confirmed in clones LPB1, LPB3 and LPB7. e Double labeling for the IMM marker CIII-Core I and βIII-TUBULIN in DMSO- or CCCP-treated DF-iPSC- and LiPSC-derived neurons from a healthy donor or a PARK2 patient. Scale bar = 50 μm. f The ratio of the IMM area was determined by quantifying the CCCP/DMSO ratio of CIII-Core I staining in the βIII-TUBULIN-positive cells (n = 3 independent experiments; means ± SEM; **p < 0.01; Student’s t-test). g The CCCP treatment significantly decreased the ratio of TH-positive neurons in the PARK2 patient-derived neurons compared with the control (n = 3 independent experiments; means ± SEM; **p < 0.01; Student’s t-test). h, i Analysis of oxidative stress using the CellROX® Green Reagent. The PARK2 neurons showed increased CellROX® fluorescence compared with the control neurons (n = 3 independent experiments; means ± SEM; **p < 0.01, ***p < 0.001; Student’s t-test). Scale bars = 500 μm