Fig. 1
SA-β-Gal activity is inhibited by rapamycin in endothelial cells (ECs) subjected to SIPS. a Schedule of SIPS-induction, rapamycin treatment and EC culture. I: rapamycin pretreatment before H2O2 treatment, II: 6 h rapamycin treatment, III: 24 h rapamycin treatment, IV: 72 h rapamycin treatment. In II-IV experiments, rapamycin was added 2 h after H2O2 treatment. b Human coronary artery endothelial cells (HCAECs) at 72 h in (a) were stained for SA-β-Gal activity. Representative images of staining for SA-β-Gal and DAPI are shown. c SA-β-Gal-positive cells in (b) were quantitated as a percentage of total cell numbers. d Real-time PCR analysis of p16INK4a using cDNA derived from HCAECs at 72 h in (a). The results are shown after normalization against values obtained for control HCAECs (value = 1). Results are presented as means ± SD from three independent experiments. e Cell cycle analysis of HCAECs at 72 h in (a). Representative data are shown. ap < 0.05 vs. the Ctr and bp < 0.05 vs. the H2O2-treated HCAECs. Ctr Control (Ctr): untreated cells. SIPS, stress-induced premature senescence