Figure 4: Effect of HMGB1 Overexpression on DNA Damage and DDR After Ang II Stimulation

(A, B) Representative images and analysis of immunostaining of nuclear foci of γ-H2AX (green), α-actinin (red), and DAPI (blue) of neonatal rat CMs (NRCMs) transfected with pcDNA-HMGB1 or control vector and then treated with saline or Ang II for 24 h. The foci number of γ-H2AX per NRCM was counted. Scale bars = 20 μm. n = 6 per group. (C, D) Representative images and analysis of Western blots of γ-H2AX. NRCMs transfected with pcDNA-HMGB1 or control saline and then treated with vehicle or Ang II for 24 h (n = 5 per group). (E, F) Representative images and analysis of immunostaining of nuclear foci of p-ATM (red), α-actinin (green), and DAPI (blue) of NRCMs transfected with pcDNA-HMGB1 or control vector and then treated with saline or Ang II for 24 h. The foci number of p-ATM per NRCM was counted. Scale bars = 20 μm. n = 5 in control vector saline group; n = 5 in control vector Ang II group; n = 4 in pcDNA-HMGB1 saline group; n = 5 in pcDNA-HMGB1 Ang II group. (G, H) Representative images and analysis of Western blots of p-ATM and total ATM. NRCMs transfected with pcDNA-HMGB1 or control vector and then treated with saline or Ang II for 24 h (n = 4 per group). ∗∗p < 0.01; ∗∗∗p < 0.001. Abbreviations as in Figures 1, 2, and 3.