Fig. 6.

Concurrent down-modulation of GRK2, AKT, and Pin1 protein levels in early stages of reperfusion in porcine hearts. Samples from non-ischemic (controls) or from the ischemic area at risk of lysates obtained from porcine hearts after 40 min of ischemia and 5 or 120 min of reperfusion as detailed in Methods were analyzed with specific antibodies for GRK2 (A), total AKT (B) AKT phosphorylated at Ser473 (C) or total Pin1 levels (D). Data were normalized by GAPDH (panels A, B, D) or total AKT (panel C) loading. (E) Global activity of AKT towards its substrates was assessed in pig cardiac lysates from the indicated conditions by dot-blot, using a pan phospho-AKT substrate-specific antibody, and data normalized by GAPDH loading. In all panels, normalized data were represented as fold-change with respect to control situation (mean value of myocardial control area at 5 min of I/R) and are mean ± SEM, n = 3–5 pigs per condition. *p < .05; **p < .01, ***p < .001 compared with the conditions indicated with the lines [1-way ANOVA and Tukey's post hoc test]. Representative blots are shown.