Fig. 2. The silencing of PPAR suppressed the senescence-associated features and cell proliferation in 2BS and WI-38 cells. 2BS and WI-38 cells transfected with the expression plasmids pSilencer 2.1-U6 neo (RNAi vector) or pSilencer-PPAR (siPPAR) were analyzed for the relative senescence markers (all transformants of 2BS at PD51 and WI-38 at PD47). Cells were treated with 20 µM troglitazone or DMSO (vehicle) as indicated. (A) Western blot analysis of PPAR silencing in siPPAR-transfected cells compared with RNAi-vector-transfected cells. Western blotting was performed using specific antibodies against PPAR as indicated. The β-actin lane serves as a loading control. (B) RNAi-vector-transfected and siPPAR-transfected cells were stained for SA-β-gal activity (blue), a classical marker of senescence. (C) Growth curves of RNAi-vector-transfected and siPPAR-transfected 2BS cells were determined by the MTT assay. Values are the mean ± s.d. of triplicate points from a representative experiment (n=3), which was repeated three times with similar results. Values accompanied by different symbols are statistically significantly different from each other. (D) Flow-cytometry analysis of RNAi-vector-transfected and siPPAR-transfected 2BS cells. Each experiment was performed at least three times. The table shows the representative data. The graph depicts data from three independent experiments (means ± s.d.). ^*P0.05 vs vehicle-treated RNAi-vector cells.