Figure 6.

Overexpression of Gadd45α protects H9C2 rat myocytes against hydrogen peroxide (H₂O₂) and recombinant Retn-induced DDR. H9C2 cells were transfected with Gadd45α or empty control by Retroviral Gene Transfer and Expression system, then treated with 200 μM H₂O₂ for 24 h or 100 ng/mL recombinant Retn for 48 h. (A) Real-time PCR analysis of the expression levels of Gadd45α in Gadd45α overexpressing H9C2 cells vs. control (n = 3). (B) Expression levels of Parp1 and Xrcc1 were analysed by real-time PCR (n = 3). (C and D) Representative images and analysis of Western blots of γH2AX (n = 3). Protein levels were normalized to GAPDH, a loading control; mRNA levels were normalized to 18S. (E and F). Representative images and analysis of immunostaining of nuclear foci of pATM (green) and DAPI (blue) (n = 6, scale bar, 100 µm). Values are shown as mean± S.E.M. with each experiment performed in biological and technical replicates as indicated. Significance was evaluated by Student’s t-test or one-way ANOVA with Tukey’s post-hoc test. P values <0.05 were considered significant.