Figure 7
T3/mH2O2 signaling inhibits the activation of cell cycle checkpoint proteins by increasing CREB-mediated Wip1 expression. (A) Representative immunoblots of lysates from cultured neonatal cardiomyocytes, and their quantitative analyses, showing that T3 inhibits ATM, CHK2 and p53 phosphorylation, while increasing CREB phosphorylation and Wip1 phosphatase expression. H2O2 scavenging, by PEG-catalase, or IGF-1 inhibition, using a IGF-1 neutralizing antibody (IGF-1 Neut Ab), inhibited these effects of T3. ***P < 0.001, compared to vehicle controls. (B) Representative immunoblot, and their quantitative analyses, showing that knockdown of CREB using siRNA prevents T3-dependent Wip1 expression in cardiomyocytes. ***P < 0.001, compared to T3 + Scrambled siRNA controls. (C) Representative immunoblot, and their quantitative analyses, showing that knockdown of Wip1 using siRNA prevents T3 from inhibiting ATM, CHK2, and p53 phosphorylation, but not stimulation of IGF-1 expression. ***P < 0.001, comparing T3-treated groups to vehicle-treated groups for cardiomyocytes pretreated with scrambled siRNA or Wip1 siRNA. †††P < 0.001, comparing the effects of Scrambled (Sc) siRNA or Wip1 siRNA pretreatment on cardiomyocytes exposed to either vehicle or T3 treatment. In (A–C), immunoblots are representative of four biological replicates. Error bars indicate SEM. n = 4/group.
