PCAF Accelerates Vascular Senescence via the Hippo Signaling Pathway

Figure 4

PCAF mediated PA-induced endothelial senescence through Hippo signaling pathway. (a) KEGG analysis showed that PCAF may regulate endothelial senescence via Hippo-YAP pathway. (b) Western blotting analysis and qualification of phosphorylated YAP expression in HUVECs treated with PA for the indicated time points. (c) Immunofluorescence of YAP localization in HUVECs treated with or without PA. Scar bars, 10 μm. (d) Expression of total YAP proteins was analyzed by immunoblot in nuclear and cytoplasmic protein extractions from in HUVECs treated with or without PA. (e) and (f): HUVECs were transfected with YAP siRNA or overexpression plasmid. The expression of p53, p16, pH2A.X and p21 protein were by immunoblotting. (g) and (h) HUVECs were transfected with YAP siRNA or overexpression plasmid, IL-6 and IL-1β mRNA expression were analyzed by RT-PCR. (i) IHC staining and its qualification of vessels from 4w and one and half-year-old mice with YAP antibody. Scale bars, 100 μm. (j) and (k) HUVECs were transfected with PCAF siRNA or overexpression plasmid, the expression of p-YAP and t-YAP protein were by immunoblotting. (l) HUVECs were transfected with PCAF siRNA or overexpression plasmid treated with or without PA. Expression of YAP proteins was analyzed by immunoblot in nuclear and cytoplasmic protein extractions. (m) Immunofluorescence of YAP localization in HUVECs infected with si PCAF or PCAF plasmid or vector. Scar bars, 10 μm. (n) and (o) Flag-tagged YAP or Flag-tagged PCAF was transfected into HUVECs before stimulated with PBS or PA for 48 h, Co-IP were performed by using corresponding magnetic beads. t-YAP: total YAP; p-YAP: phosphorylated YAP. Data are expressed as ( for each experiment. , , , n.s. ).

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