FIGURE 5.

Accumulation of p21 protein causes G₂ arrest of cell cycle in cardiomyocytes. A, down-regulation of p21 in D1NLS/CDK4 cardiomyocytes infected with adenovirus encoding siRNA specific for p21. At 48 h post-infection, the cells were harvested and extracted for CDK2 and CDC2 kinase assay. B, measurement of cell number of cardiomyocytes infected with adenoviruses for D1NLS/CDK4 and p21 siRNA. At each day indicated, cell number was counted, and the relative cell proliferation was expressed as mean ± S.E. of three independent experiments. C, cell cycle analysis of cardiomyocytes infected with a combination of adenoviruses for D1NLS/CDK4, CDK2, and p21 siRNA as indicated using a laser scanning cytometer. D, quantitative measurement of H3P-positive cardiomyocytes. Cells were treated as in C. At 48 h post-infection, cells were fixed and immunostained with anti-phospho-histone H3 (H3P) and anti-tropomyosin antibodies. Cardiomyocytes positive for H3P were counted and expressed as the percent of total. Data are the means with S.E. of three independent experiments.