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. 2016 Jul 21;6:29994. doi: 10.1038/srep29994

Figure 6. The hAFS secretome antagonizes Dox-elicited damage of human CPC.

Figure 6

(a) Representative images of human c-kit+ CPC (hCPC) stained for senescence associated (SA) β-galactosidase: untreated cells (Ctrl), cells exposed to 0.2 μM Dox (Dox), and cells incubated with 40 μg/ml of the hAFS-CMHypo (hAFS-CMHypo + Dox) or with the hNCTC-CMHypo (hNCTC-CMNormo + Dox) prior to Dox treatment. Scale bar 100 μm. (b) Percentage of hCPC expressing SA β-galactosidase (mean ± s.e.m.). Ctrl: 14.8 ± 2.7%, Dox: 50.1 ± 6.4%, hAFS-CMHypo + Dox: 24.6 ± 1.9%, and hNCTC-CMHypo + Dox: 47.5 ± 7.0%; *p < 0.05 (p = 0.0285). (c) Representative western blot for the expression of the senescence marker p16INK4a (17kDa; housekeeping GAPDH, 37kDa) after exposure to 0.2 μM Dox with or without pre-incubation with 40 μg/ml of the hAFS-CMHypo or with hNCTC-CMHypo. Full-length blots are represented in Supplementary Fig. S4. (d) Densitometry of the western blot for p16INK4a with same conditions as in (c). (e) Representative images of hCPC stained for cleaved-caspase-3 (Caspase-3): untreated cells (Ctrl), cells exposed to 1 μM Dox (Dox), and cells incubated with 40 μg/ml of the hAFS-CMHypo (hAFS-CMHypo + Dox) or with the hNCTC-CMHypo (hNCTC-CMNormo + Dox) prior to Dox treatment. Scale bar 100 μm. (f) Percentage of hCPC expressing cleaved caspase-3 (% Caspase-3+ cells, mean ± s.e.m.). Ctrl: 0.90 ± 0.1%, Dox: 4.5 ± 0.6%, hAFS-CMHypo + Dox: 3.8 ± 0.5%, and hNCTC-CMHypo + Dox: 6.4 ± 1.5%.