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. 2011 May 30;193(5):867–884. doi: 10.1083/jcb.201010051

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© 2011 Heo et al.

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Figure 5. — ONOO⁻ mediates d-flow–induced PKCζ activation, p53 SUMOylation, and EC apoptosis. (A) ONOO⁻ mediates d-flow–induced PKCζ activation and p53 SUMOylation. HUVECs were pretreated by 5 µM ebselen, 20 µM L-NAME, and 10 µM Mn-TBAP for 30 min and exposed to d-flow for 3 h. PKCζ phosphorylation at Thr560 and p53 SUMOylation were determined as described in Materials and methods. (B and C) Densitometry analyses of p53 SUMOylation (B) and PKCζ phosphorylation (C) were performed as described in Fig. 1. **, P < 0.01 compared with the vehicle control in static condition, and ^#, P < 0.01 compared with the vehicle control in d-flow stimulation for 3 h. (D and E) HUVECs were pretreated by each inhibitor for 30 min and exposed to d-flow for 36 h followed by TUNEL staining as described in Materials and methods (D), and quantification of apoptosis is shown as the percentage of TUNEL-positive cells (E). Bars, 30 µm. Data are from three separate experiments using two or more different EC preparations (**, P < 0.01 compared with the vehicle control in static condition, and ^#, P<0.05 and ^##, P<0.01 compared with the vehicle control in d-flow stimulation for 36 h). Error bars show means ± SD. Molecular masses are given in kilodaltons. IB, immunoblot. IP, immunoprecipitation.