Higher Cell Seeding Density Decreases Cell Cycle Progression.

<p>(A) A representative histogram (left) of low density (1.3 x 10⁴ cells/cm², black line) and high density (5.3 x 10⁴ cells/cm², red line) seeded CA1S hESCs stained for DNA content by propidium iodide to indicate cell cycle state within the depicted gates 24-hours after seeding. (B) Single cells gated for uniform DNA width were assessed in triplicate and quantified as either G0/G1, S or G2/M phases using the gates in (A) as a percentage of the total single cell population. Four cell seeding densities of CA1S cells (1.3, 2.6, 3.9 and 5.3 x 10⁴ cells/cm²) along with 5.3 x 10⁴ cells/cm² seeded cells treated overnight with 2% DMSO to induce cell cycle arrest (2% DMSO) were quantified. * represents significant difference from 1.3 x 10⁴ cells/cm² by one-way ANOVA with Bonferroni post-hoc test within the same cell cycle population. (C) Representative images and quantification of immunocytochemistry of pRb S780 (green, nuclei are blue). pRb S780 positive mitotic cells were quantified as a percentage of the total cell populations in five randomly selected images. Different superscripts (a, b, c) are significantly different from each other by one-way ANOVA with Bonferroni post-hoc test. Scale bars are 100 μm.