Figure 2
p62 deficiency disrupts the NADH homeostasis. (A) Time-course representative traces of NADH autofluorescence from untransfected, SCR and p62 KD in SH-SY5Y cells. The uncoupler FCCP (1 μM) maximises mitochondrial respiration and therefore minimises mitochondrial NADH (0%). NaCN (1 mM) was then added to block mitochondrial respiration and thereby maximise mitochondrial NADH (100%). The traces represent the mean of at least 20 cells on a single coverslip ±SEM. (B) Graphical description of the NADH homeostasis analysis by monitoring the NADH autofluorescence in cells. NADH redox indexes were obtained by calculating the initial NADH autofluorescence when the minimum NADH autofluorescence is normalised to 0% and the maximum to 100%. Both the NADH redox index (the initial redox level expressed as percentage of the range) and the NADH pool are described graphically. (C,D) NADH redox indexes from untransfected, SCR and p62 KD SH-SY5Y cells (C) and control and p62 deficient fibroblasts (D) representing the mean of at least 3 independent experiments ±SEM. In all cases ** indicates p < 0.01 and ***indicates p < 0.001 compared with the values in control cells. (E,F) The NADH pool was expressed as absolute values between maximal and minimal respiration in untransfected, SCR and p62 KD SH-SY5Y cells (E) and control and p62 mutant fibroblasts (F). Data represent the mean of at least 3 independent experiments ±SEM. In all cases **indicates p < 0.01 and ***indicates p < 0.001 compared with the values in control cells.
