Fig. 8

RGS6 overexpression reduced apoptosis in MLE-12 cells. A Immunofluorescent staining showed RGS6 positional changes after LPS stimulation. Cell nucleus was stained by DAPI (blue fluorescence). Cytoplasm was stained by anti-β Tubulin antibody (green fluorescence). RGS6 protein was stained by anti-RGS6 antibody (red fluorescence). Scale bar: 20 μm. B RGS6 overexpression efficiency in MLE-12 cells was shown by qRT-PCR. Control group was flagged as NC, while RGS6 overexpression group was flagged as OE. C Flow cytometry showed Annexin V-FITC/PI changes in different groups. D Quantitative analysis showed the percentage of early apoptotic cells (FITC + PE−). E Bcl-2 mRNA relative expression levels in different groups. F Western blotting showed protein expression levels in different groups. G Caspase 3 activity assays for live cells in different groups. Cell nucleus was stained by Hoechst 33342 (blue fluorescence). Scale bar: 100 μm. NC-C, MLE-12 cells transfected with control plasmid for 24 h followed by culturing for another 24 h. OE-C, MLE-12 cells transfected with RGS6 overexpression plasmid for 24 h followed by culturing for another 24 h. NC-LPS, MLE-12 cells transfected with control plasmid for 24 h followed by stimulating with LPS (1 μg/ml) for another 24 h. OE-LPS, MLE-12 cells transfected with RGS6 overexpression plasmid for 24 h followed by stimulating with LPS (1 μg/ml) for another 24 h. Data are shown as mean ± SEM of triplicate. *P < 0.05 vs. NC-C group; **P < 0.01 vs. NC-C group; ***P < 0.001 vs. NC-C group. ^#P < 0.05 vs. NC-LPS group; ^##P < 0.01 vs. NC-LPS group. ^###P < 0.001 vs. NC-LPS group