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. 2023 Mar 30;66(7):1306–1321. doi: 10.1007/s00125-023-05905-8

Fig. 7.

Fig. 7

Exenatide protects Wolfram syndrome iPSC-derived neural precursors and cerebellar neurons. iPSCs Wolf 2010-9.4, Wolf 2010-11.1 and Wolf-2010-07.1 were differentiated into cerebellar neuron-like cells. (a) Differentiation timeline with representative phase contrast images of iPSCs (scale bar, 120 μm), neural rosettes, NPCs and cerebellar neuron-like cells (scale bars, 50 μm). (b) Representative immunofluorescence images for key markers OCT4, nestin, vimentin, PAX6, β-tubulin III, synaptophysin, GFAP, KIRRE2, ZIC1 and calbindin at day 12 of the first differentiation, in early NPCs (passage 1, P1) and at days 10 and 21 of the second differentiation. Scale bars, 30 μm. (c) Mitochondrial function by Seahorse in NPCs (n=3) and cerebellar neuron-like cells (n=3) exposed or not for 72 h to exenatide (500 nmol/l) or forskolin (20 μmol/l). Mitochondrial respiration was measured basally and after sequential injection of 20 mmol/l glucose, ATP synthase inhibitor oligomycin (5 μmol/l), oxidative phosphorylation uncoupler FCCP (4 μmol/l), and electron transport chain inhibitors rotenone and antimycin (1 μmol/l). Results are shown as means ± SEM of the two cell types together. (d, e) WFS1 iPSC-derived NPCs (squares) and cerebellar neuron-like cells (triangles) were exposed or not (CT) for 48 h to tunicamycin, alone or together with exenatide or forskolin. (d) Oxidative stress was measured by HPF. Menadione-treated cells were used as a positive control. (e) Apoptosis was determined by Hoechst 33342/propidium iodide. Extremities of floating bars are maximal and minimal values; horizontal line shows median. Symbols with the same colour come from one differentiation. *p<0.05, **p<0.01, ***p<0.001 Ex vs CT, †p<0.05, †††p<0.001 treated vs CT; §§p<0.01, §§§p<0.001 Tuni vs Tuni+Ex or Tuni+Fk; p<0.05 Ex vs CT, by two-way or one-way ANOVA followed by Dunnet or Holm–Sidak correction for multiple comparisons. CT, control (vehicle); Ex, exenatide; Fk, forskolin; GFAP, glial fibrillary acidic protein; P, passage; PAX6, paired box protein 6; Pos, positive control; Tuni, tunicamycin