Fig. 6

Knockdown of NR3C1 activates ER stress and mitophagy in ccRCC. TEM of ER stress in the control and sh-NR3C1 groups of ACHN (A) and 786-O (B) cells. Shown are representative images, with arrows indicating ER stress. (Scale bar =1.0 μm; enlarged scale bar =0.4 μm). C Expression levels of ER stress pathway proteins in the two groups of ACHN and 786-O cells at different times of GC treatment using Western blot. (GC=1200 nM). TEM of mitophagy in the control and sh-NR3C1 groups of ACHN (D) and 786-O (E) cells. Shown are representative images, with arrows indicating mitochondrial autophagosomes. (Scale bar =1.0 μm; enlarged scale bar =0.4 μm). F Bar graphs represent the statistical results of mitochondrial membrane potential detection. Representative fluorescence images of mitochondrial membrane potential in the control and sh-NR3C1 groups of ACHN (G) and 786-O (H) cells were stained by the JC-1 probe, with CCCP used as a positive control. (Scale bar =100 μm). I Colocalization of lysosomes and mitochondria in ACHN cells after NR3C1 knockdown was determined by confocal microscopy. Mitochondria and lysosomes were labeled with mitotracker and lysotracker, respectively. (Scale bar =10 μm). J The overlap coefficient of immunofluorescence co-localization. Data presented as mean ± SD with three replicates. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001. TEM, transmission electron microscopy; GC, glucocorticoids; CCCP, carbonylcyanide-m-chlorophenylhydrazone; Mito-tracker, green fluorescent probe of the mitochondria; Lyso-tracker, red fluorescent probe of lysosome