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. 2019 Oct 5;48:605–618. doi: 10.1016/j.ebiom.2019.09.019

Fig. 1.

Fig. 1

Ischemia, reperfusion and conditioning treatments trigger marked and dynamic changes in cardiac GRK2 protein levels and phosphorylation status. Isolated rat hearts were exposed to 40 min of ischemia alone 40or with preconditioning I40PreCo(I) followed by reperfusion for the times indicated (I/R) in the presence or absence of preconditioning (I/R-PreCo) or post-conditioning (I/R-PosCo) interventions as detailed in Methods. Control group (Nx, normoxia condition) was exposed to60 min of continuous perfusion without ischemia. (A) GRK2 protein levels were analyzed by western-blot with a specific total anti-GRK2 antibody as detailed in Methods. GAPDH expression was used as loading control. (B–C) The extent of GRK2 phosphorylation on different residues (Ser670 or Ser685) was monitored in cardiac lysates by western-blot using phosphosite-specific antibodies. Data were normalized to total GRK2 levels GAPDH expression was used as loading control. In all panels, data are mean ± SEM, n = 3–4 rats per condition. In all panels, data were analyzed by comparing the different experimental situations (I/R, I/R-PreCo and I/R-PosCo) to the normoxic condition [2-way ANOVA followed by Bonferroni's post-hoc test, †p < .05; ††p < .01, †††p < .001]. In addition, we compared I/R pre-Co and I/R post-Co conditions versus I/R alone (*p < .05; ***p < .001) or between conditioning situations (#p < .05; ###p < .001) [1-way ANOVA and Tukey's post hoc test]. Representative blots are shown.