Fig. 2.

Neonatal ventricular rat myocytes (NVRM) were plated (48 h) on collagen type I-coated (0.1 mg/ml) and fibronectin-coated (0.05 mg/ml) polyacrylamide (PA) gels of varying stiffnesses. A and B: on soft gels (shear modulus G ∼100 Pa, 300 Pa), cells appeared rounded, with poor sarcomere organization. C and D: at physiological stiffness (G ∼5 kPa, 10 kPa), cells exhibited striated F-actin-stained (red) and α-actinin-stained (green) fibers and increased spread area while maintaining a high aspect ratio. E and F: on a stiff surface (G ∼30 kPa, glass), cells exhibited prominent F-actin filaments devoid of striations, consistent with a stress fiber-like appearance. Scale bars = 10 μm for all images. G: comparative bar graph of cell-spreading area on extracellular matrices (ECM) of varying stiffnesses reveals an increase in spread area as a function of increasing stiffness. The spread areas of myocytes showed a statistically significant increase with increasing substrate stiffness from 0.1 kPa to glass. H: aspect ratio of cardiac myocytes responsive to varying stiffnesses. Significant differences in aspect ratio were observed between 0.1 kPa and 0.3 kPa. The aspect ratio was significantly higher for 10 kPa (physiological stiffness) compared with 0.3 kPa, 5 kPa, and 30 kPa. Group comparisons were made using ANOVA, and significance was determined using the t-test between two substrates of different stiffness (e.g., 0.3 kPa vs. 5 kPa, pairs shown in brackets). *P < 0.05; **P < 0.01. Number of samples is shown inside each vertical column (n = 199–361). Error bars indicate standard error.