Figure 2

Effect of 3BDO on oxLDL-induced autophagy in HUVECs.

(a), Western blot analysis of ATG13 and p-ATG13 protein level and quantification (cropped, full-length blots are in Supplementary Figure S7), with nLDL, 50 μg/ml; oxLDL, 50 μg/ml; 3BDO-L, 60 μM; and 3BDO-H, 120 μM, for 12 h. nLDL control data were set to 1. Data are mean ± SEM; ATG13, *p = 0.017 vs. nLDL, #p = 0.024 vs. oxLDL, & p = 0.024 vs. oxLDL; p-ATG13/ATG13, **p = 0.004 vs. nLDL, #p = 0.039 vs. oxLDL, & p = 0.011vs. oxLDL. n = 3. (b), Western blot analysis of LC3 –II and p62 and quantification (cropped, full-length blots are in Supplementary Figure S8), with nLDL, 50 μg/ml; oxLDL, 50 μg/ml; 3BDO-L, 60 μM; and 3BDO-H, 120 μM, for 12 h. Protein levels were normalized to that of β-actin. nLDL control data were set to 1. Data are mean ± SEM; LC3–II, **p = 0.0008 vs. nLDL, #p = 0.01 vs. oxLDL, &&p = 0.0005 vs. oxLDL; p62, **p = 0.004 vs. nLDL, ##p = 0.003 vs. oxLDL, &&p = 0.009 vs. oxLDL. n = 3. (c), Immunofluorescence staining of LC3, with nLDL, 50 μg/ml; oxLDL, 50 μg/ml; 3BDO-L, 60 μM, for 12 h. Bar = 5 μM. Bar charts showed the quantification of average endogenous LC3 puncta per cell. Different fields of view (>3 regions) were analyzed on the microscope for each labeling condition and representative results are shown. Data are mean ± SEM.