Figure 1. BCL-2 family members are elevated in senescent cells and provide resistance to apoptosis.

(a) IMR-90 human fibroblasts that were induced to senesce either through DNA damage (DIS), replicative exhaustion (RS) or oncogene expression (OIS), as well as controls proliferating cells (growing, G) and empty vector-transfected (V) cells, were treated for 10 h with TNF-α and CHX (TNF-α) or with vehicle (DMSO). Cell survival relative to vehicle-treated cells was determined by quantification of the remaining adherent cells. Histograms indicate the percentages of surviving senescent (DIS, RS and OIS) cells compared with G or V controls. Data are presented as mean±s.e.m of three repeats, performed in triplicates. (b) Western blot analysis of cleaved PARP, ICAD and caspase-3 proteins from senescent and control IMR-90 cells (labelled as in a), indicating levels of apoptosis after treatment with TNF-α and CHX. (c) Percentage survival of senescent and control cells (labelled as in a) 24 h after UV irradiation (80 J m⁻²). Data are presented as mean±s.e.m of three repeats, performed in triplicates. (d) Expression of BCL-2 family members (BCL-W, BCL-XL, BCL-2 and MCL-1) and of senescence effector proteins (p16, p21, p53) in senescent (DIS, RS, and OIS) and control (G, V) cells of both human (IMR-90) and mouse (MEF) origin. (e) Percentage survival of senescent and control cells (as in a) as well as in quiescent (Q) control cells after treatment for 24 h with the indicated concentrations of ABT-737, an inhibitor of BCL-W, BCL-XL and BCL-2. Data are presented as mean±s.e.m of three repeats, performed in triplicates. (f) Percentage survival of senescent (DIS and OIS) and control (G and V) cells treated with ABT-737 (10 μM) for 24 h with or without pretreatment for 6 h with the pan-caspase inhibitor z-VAD-fmk. Data are presented as mean±s.e.m of three repeats, performed in triplicates. (g) Western blot analysis of cleaved PARP and caspase-3 in the samples described in f. Data in b,d,g are representative blots of at least two independent experiments each. Data were analysed using Student's t-test. *P<0.05, **P<0.005.