Figure 5. Effect of ROCK inhibitors on collagen gel contraction. Cells grown in collagen gels were incubated without (control) or with ROCK inhibitors Y-27632 (5 μM or 10 μM), and Y-39983 (1 μM or 5 μM) for 48 h. Changes in the area of the gels were calculated using the ImageJ program from NIH. The initial area of the gels (measured before the addition of drugs) was ~140 mm². A: The presence of serum contracted the gels significantly. This serum-induced contraction was opposed by both Y-27632 and Y-39983, with the latter having a more significant effect. Blebbistatin (10 μM), a specific myosin II ATPase inhibitor, was employed to confirm that the observed changes in the gel area resulted from increased actomyosin contraction. B: A bar graph of the results from several independent experiments. As can be noted, the absence of serum led to a lesser contraction (by 18±0.5%) of the gels compared to serum (~60 mm²) after 48 h. The ROCK inhibitors significantly opposed the decrease in the gel area. At 10 μM Y-27632, the gel area had increased by 50%, while, even at 1 μM, Y-39983 opposed actomyosin contraction more effectively (86±5%) compared with the control. The gel area obtained with 10 μM of blebbistatin (denoted as Bstat) was similar to that obtained with 5 μM Y-39983. The data are represented as mean±SEM.