Figure 8.

The inhibition of CDK4 or CDK7 by kinase inhibitors blocks autophagy activation induced by SiNPs. (A) GFP-LC3 NRK cells were incubated with the inhibitor ON123300 for 24 h, and then treated with 16-nm SiNP at 60 μg/mL for 0, 12, 16, 20 or 24 h. Scale bar: 10 μm. (B) The GFP-LC3 puncta were observed and measured by confocal microscopy, and the number of GFP-LC3 puncta was quantified in ≥ 30 cells. (C) The immunoblotting analysis of the LC3-II expression in NRK cells, which were treated with ON123300 or DMSO for 24 h and then stimulated by 16-nm SiNPs. The relative expression of (D) LC3-II, (E) CDK4 or (F) p-CDK4 was calculated by the ratio of LC3-II:ACTB, CDK4:ACTB or p-CDK4:ACTB, respectively. (G) GFP-LC3 NRK cells were incubated with the inhibitor THZ1 for 24 h and then treated with 16-nm SiNP at 60 μg/mL for 0, 12, 16, 20 or 24 h. Scale bar: 10 μm. (H) The GFP-LC3 puncta were observed and measured by confocal microscopy, and the number of GFP-LC3 puncta was quantified in ≥ 30 cells. (I) The immunoblotting analysis of LC3-II in NRK cells, which were treated with THZ1 or DMSO and then stimulated by 16-nm SiNPs. The relative expression of (J) LC3-II, (K) CDK7 or (L) p-CDK7 was calculated by the ratio of LC3-II:ACTB, CDK7:ACTB or p-CDK7:ACTB, respectively. * p < 0.05; *** p < 0.001