Figure 4: Vacuoles have reference to autophagy. (a) MDC staining of autophagosomes after 20 μM Rottlerin treatment for 6, 18, and 24 h; o.m.: original magnification; (b) western blotting analysis of the autophagosome marker LC3I-II after 20 μM Rottlerin treatment for 1–24 h and increased LC3-II/LC3-I ratio, calculated by densitometry; (c) autophagic clearance of SQSTM1/p62. β-actin was used as loading control. Representative of three independent experiments.