Figure 1.

AGE-BSA-induced SMAD3 activation is associated with autophagic blockage in HK-2 cells. (A) Double immunofluorescence staining of SQSTM1 and TGFB1 in HK-2 cells exposed to 30 μg/ml AGE-BSA or 30 μg/ml Co-BSA for 24 h. (B and C) Quantification of SQSTM1 puncta and TGFB1 fluorescence intensity. (D) Double immunofluorescence staining of SQSTM1 and phospho-SMAD3 in AGE-BSA or Co-BSA-treated HK-2 cells. (E and F) Quantification of SQSTM1 puncta and phospho-SMAD3 protein-positive nuclei. (G–K) Western blot analysis of LC3, SQSTM1, and phospho-SMAD3 expression in AGE-BSA or Co-BSA-treated HK-2 cells. Bars represent means ± SEM from at least 3 independent experiments. *P< 0.05, **P< 0.01 and ***P< 0.001. DAPI was used to stain nuclei. Scale bar: 10 µm