Reduced SQSTM1 expression increases ROS levels and senescence of VSMCs. RASMs were transfected with siRNA control or siSqstm1 for measurements of SQSTM1 and CDKN1A expression (A-C), and SA-GLB1 activity, using FDG, in response to 100 nM AGT II or 50 μM zinc for 3 d (D). ppargc1a−/- VSMCs were infected with 1 × 107 PFU of adenovirus control (Ad.Control) or adenovirus containing human SQSTM1 gene (Ad.SQSTM1) for ROS measurements (E) and SA-GLB1 activity using FDG (F). Ppargc1a+/+ and ppargc1a−/- VSMCs infected with Ad.Control or Ad.SQSTM1 were lysed for analysis of SQSTM1 and LC3-II expression. (G-I). Expression was expressed as fold change compared with siControl-transfected cells. Aortas of 2-month-old mice Sqstm1+/+, Sqstm1± and sqstm1−/- in the C57Bl/6 background (8 males each) were cleaned, fixed and tested for SA-GLB1 activity using FDG (J). Fluorescence was adjusted by the weight of aortas. * denotes p < 0.01. Body weight for each genotype is expressed in grams (g) (K). Aortas of each genotype (2-month-old, males) were cleaned of periadventitial fat, lysed and 80 μg of total extracts were separated in a 4–20% precast Criterion gels (L). Quantification of TRP53 and LC3-II adjusted by ACTB is shown in M and N, respectively. * denotes p < 0.05.