Fig. 2: PTEN regulates autophagy activity followed by ROS level and VNR-induced apoptosis in AS2 and A549 cells.

a A representative western blot analysis showing the expression of Akt, p-Akt, Keap1 and PTEN in the AS2 cells without or with the transfection of plasmids containing GFP-PTEN. GFP was used as a vector control. β-actin was used as an internal control. The relative densities of the measured protein bands are also shown. b A representative western blot analysis showing the expression of Akt, p-Akt, Keap1, PTEN in the A549 cells with shLuc and shPTEN. c CM-H₂DCFDA staining, followed by a flow cytometric analysis, was used to determine the levels of ROS in AS2 cells transduced with GFP and GFP-PTEN. For the flow cytometric analyses, the percentages are the means ± SDs of three individual experiments. ***P < 0.001. d CM-H2DCFDA staining, followed by a flow cytometric analysis, was used to determine the levels of ROS in A549 cells with shLuc and shPTEN. For the flow cytometric analyses, the percentages are the means ± SDs of three individual experiments. **P < 0.01. e Nuclear PI staining and subsequent flow cytometric analysis determined cell apoptosis in VNR-treated AS2 cells transduced with GFP and GFP-PTEN. The percentages (%) of apoptotic cells are shown as the means ± SDs of three individual experiments. **P < 0.01. f Nuclear PI staining and subsequent flow cytometric analysis determined cell apoptosis in the VNR-treated A549 cells with shLuc and shPTEN. The percentages (%) of apoptotic cells are shown as the means ± SDs of three individual experiments. *P < 0.05. A representative western blot analysis showing the expression of p62 (g) and LC3 I and II (h) in AS2 cells transduced with GFP and GFP-PTEN. β-actin was used as an internal control. The relative densities of the measured protein bands are also shown. A representative western blot analysis showing the expression of p62 (i) and LC3 I and II (j) in the A549 cells with shLuc and shPTEN