Additional file 2 of Human midbrain organoids reveal the characteristics of axonal mitochondria specific to dopaminergic neurons

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Additional file2. Analysis of mitochondrial membrane potential in neurites derived from other TH-GFP iPSC lines. (a) Live-cell imaging of neurites of dopaminergic and non-dopaminergic neurons derived from other TH-GFP iPSC lines. Green and white dotted lines indicate mitochondria in GFP-positive and GFP-negative neurites, respectively. “PRKN” represents PRKN-mutant patient. Scale bar, 2 µm. (b) Mean TMRM MFI in all mitochondria per neurite from the live-cell images. Data were acquired from GFP-negative (n = 20) and GFP-positive (n = 12) neurites from two fields in the control 201B7 T1-3 line, from GFP-negative (n = 19) and GFP-positive (n = 6) neurites from two fields in the control WD39 T1-2 line, and from GFP-negative (n = 16) and GFP-positive (n = 8) neurites from two fields in the PRKN-mutant PB2 T1-1 line. The graph shows the relative comparison of the mean TMRM MFI of mitochondria in GFP-negative and GFP-positive neurites. “PRKN” represents PRKN-mutant patient. Values are shown as the mean ± SEM. The differences were evaluated using the nonparametric Mann–Whitney U test. **P < 0.01, ***P < 0.001. The mean TMRM MFI of mitochondria significantly differs between dopaminergic and non-dopaminergic neurons in the control lines B7 T1-3 (P = 0.0126; effect size, 0.383; 95% CI, 0.185 to 0.592) and WD39 T1-2 (P = 0.0002; effect size, 0.585; 95% CI, 0.336 to 0.816), whereas no significant difference is observable in the PRKN-mutant PB2 T1-1 line (P = 0.569; effect size, 0.0783; 95% CI, −0.156 to 0.419). BF: bright field, CI: confidence interval, GFP: green fluorescence protein, iPSC: induced pluripotent stem cell, MFI: mean fluorescence intensity, n.s.: not significant, SEM: standard error of the mean, TH: tyrosine hydroxylase, TMRM: tetramethylrhodamine methyl ester

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