Fig. 4.

RGS11 regulates ATF3 in cardiomyocytes. (A) Representative cardiac staining for ATF3 in a control or chemotherapy exposed patients [n = 10, scale bar = 100 μm] and correlation between ATF3 histoscores and fibrosis or cardiac troponin levels in control or chemotherapy exposed patients (n = 10). (B) Immunoblotting for ATF3 and NRG1 protein expression in heart tissue from chemotherapy exposed patients with or without fibrosis or matched controls (n = 4–5). (C–E) Control or RGS11 CRISPR KO AC-16 cells were treated with doxorubicin (3 μM, 16 h) ± ATF3 shRNA for 12 h. (C) CM-H₂-DCFDA fluorescence (total ROS; n = 5). (D) Apoptosis (cytoplasmic histone-associated DNA fragments; n = 5). (E) Protein immunoreactivity was determined via western blotting (n = 6). (F–I) Control or RGS11 KO AC-16 cells were treated with doxorubicin (3 μM, 16 h) ± pre-treatment with ATF3 shRNA for 12 h or Nox blocker DPI (1 μM) or pan-ErbB blocker cI-1033 (2 μM). (F) CM-H₂-DCFDA fluorescence (total ROS; n = 5). (G–H) NRG1 concentration in cell culture media (n = 5). (I) Apoptosis (cytoplasmic histone-associated DNA fragments; n = 5). β-Actin serves as a loading control for western blots. Immunoblots are accompanied by a densitometric quantification wherein expression is normalized to the corresponding control group. Data were analyzed by student's t-test or one- or two-way ANOVA with Sidak's post-hoc test. *P < 0.05, **P < 0.01,***P < 0.001, ****P < 0.0001. ns = not significant. Data are presented as mean ± SEM.