Fig. 6

Fibroblasts from the SPG11 patient exhibit enlarged LAMP1-positive organelles and accumulation of autolysosomes. a QPCR analysis revealed a significantly decreased expression of spatacsin mRNA in the pateint’s fibroblasts. b ICH of spatacsin (red particles, arrow indicated) in fibroblast of WT (left panel) and the SPG4 patient (right panel). c ICH analysis revealed fibroblasts from the SPG11 patient exhibit prominently enlarged LPOs. Insets in the images are enlarged (original magnification, × 9.0) to the right. d Quantification of lysosome perimeter (20 cells, > 20 lysosomes per cell). e Quantification of cells with exceed 20 enlarged LPOs (n = 3, > 30 cells per experiment). f, g Western blot analysis showed no significant difference in LAMP1 levels between the SPG11 patient and WT. h ICH of p62 (green) in fibroblast of WT (upper panel) and the SPG11 patient (lower panel). Insets in the images are enlarged (original magnification, ×9.0) to the right. i Quantification of h (50 cells). j, k Western blot analysis revealed p62 protein was significantly accumulated in the patient derived fibroblasts. l, m Western blot analysis showed the ratio of LC3-II/LC3-I was significantly increased in autophagy induced fibroblasts from the SPG11 patient. n, o Co-immunostaining with LC3 (red) and LAMP1 (green), which showed no significant difference of their overlapping between the two groups. Insets in the images are enlarged (original magnification, ×10.5) to the right. Scale bar, 10 μm. Error bars indicate SEM. *, P < 0.05. ns, not significant