Figure 1
Quantification of HCN4 and Cx40 positive cells in E11.5 mouse ventricular cell cultures following addition of exogenous ANP and or NPRA inhibitor. (A–C) Representative images of cells co-stained with sarcomeric myosin, MF20 (A) and HCN4 (B) antibodies along with Hoechst nuclear stain (C). (D–F) Cells co-stained with MF20 (D) and Cx40 (E) antibodies. (G–I) Cells were co-stained with MF20 (G) and rabbit nonimmune IgG (H) antibodies. Overlays of immunostaining signals with Hoechst staining were shown in panels C, F and I. Arrows indicate HCN4+ or Cx40+/MF20+ cardiomyocytes, arrow-heads indicate HCN4+ or Cx40+/MF20− nonmyocytes/VCS progenitor cells. A-I: scale bars = 50 μM. (J–O) Percentage distribution and ratios of HCN4+/MF20+ and HCN4+/MF20− cells (J–L) or Cx40+/MF20+ and Cx40+/MF20− cells (M–O) in E11.5 mouse ventricular cell cultures treated with ANP (0, 10, 100 and 1000 ng/ml) and or NPRA inhibitor A71915 (1 µM). N = 5 independent experiments, ~600–900 cells were counted for each group. Each bar represents mean ± SEM, *P < 0.05 Vs. 0 ng/ml (control), #P < 0.05 Vs. ANP (1000 ng or 1 µg/ml) for each panel in J–O, One-way ANOVA with Tukey’s multiple comparisons post hoc test.
