Response of SASP-RAP to senescence under different cellular contexts.

<p>(A) NRK-52E cells were treated with 4 mM 4-PBA, 20 ng/ml trichostatin A (Tri A) and 100 nM doxorubicin (Doxo) for 3 days and subjected to phase-contrast microscopy. Quantitative assessment of senescent morphology is shown in the right graph. (B) Cells were transfected with pSV40-SEAP and treated with 4-PBA, trichostatin A or doxorubicin at indicated concentrations for 3 days and subjected to chemiluminescent assay. (C) MEF were treated with 1 µg/ml etoposide for 3 days and subjected to phase-contrast microscopy and SA-β-gal staining. (D) MEF were transfected with pSV40-SEAP and exposed to 1–5 µg/ml etoposide for 3 days, and culture media were subjected to chemiluminescent assay. (E) Normal human mesangial cells (NHMC) at passage 8 (P8) and passage 13 (P13) were co-transfected with pSV40-SEAP and pEGFP-N1, treated with 1 µg/ml etoposide for 3 days and subjected to phase-contrast microscopy and SA-β-gal staining (E) and chemiluminescent assay (F). In (F), SASP-RAP activity was normalized by both transfection efficiency (percentages of EGFP-positive cells) and viable cell number, and the resultant values are shown. In reporter assays, assays were performed in quadruplicate, and data are expressed as means ± SE. Asterisks indicate statistically significant differences (<i>p</i><0.05).