Fig. 2

C9orf72 MNs have reduced lysosome numbers and altered lysosome morphology. a Fluorescent microscopy images of 40-day-old C9orf72 and isogenic control MNs labeled with SiR-Lysosome (SiR-Lyso), a dye that selectively labels Cathepsin D-positive lysosomes. Scale bar = 50 µm. b, c Quantification of the SiR-Lysosome puncta number (b) and fluorescent intensity per cell (c) from the images shown in (a); each dot represents one confocal image that was analyzed (n = 35 for all conditions). d Representative TEM images from 40-day old MNs used to quantify the size of lysosomes/late endosome (indicated by black arrowheads). Multivesicular bodies (MVB), Mitochondria (M), Golgi complex (GC), the endoplasmic reticulum (ER) and the ER whorls (ERW) are marked on the images. e, f Relative frequency distribution (e) and quantification (f) of the lysosomal circumference measured from TEM images as shown in (d); each dot represents a lysosome that was measured (C9-1, n = 197; C9-1iso, n = 472; C9-2, n = 257; C9-2iso, n = 445). Data represent mean ± SEM; data are pooled from three independent differentiations. Statistical significance was assessed by one-way ANOVA and Tukey’s multiple comparison tests (b, c) or Kruskal–Wallis test and Dunn’s multiple comparison tests (f); *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001