FIGURE 5.

siRNA targeting p65 prevents senescence mediated by the evSASP. (a) Schematic representation for the experimental settings (left panel) and timings (right panel) of the experiments performed in this figure. Reverse transfection was performed with 50 nM of the indicated siRNA in the recipient cells. After washing the siRNA out, HFFF2 recipient cells were incubated with sEV isolated from either iC or iRAS for 72 h and 3 days later several markers of senescence were determined. (b) The use of two independent siRNA targeting p65 (sip65) ‐§7 and §10‐ prevents the cell cycle arrest characteristic of evSASP senescence. A scramble siRNA control (Scr) was used as a negative control while siRNAs targeting CDKN2A (encoding for p16^INK4A) (sip16) or TP53 (sip53) were used as positive controls. (c) The percentage of cells expressing p16^INK4A protein levels and (d) stabilizing p53 protein was quantified. (e) Representative IF images for IL‐8 expression levels in HFFF2 transfected with the indicated siRNAs and treated with sEV isolated from either iC or iRAS HFFF2 cells. Scale bar: 50 µm. Data show the mean ± SEM of 6 independent experiments. Statistical analysis was performed using One‐way ANOVA. All conditions were compared to sEV from iRAS control