Fig. 4

Ectopic DNA-replication in Rb1 ^G/G ; Cdkn1b ^−/− ; E2f1 ^−/− compound mutant mice. a H&E staining of liver sections from 8-week old wild type, Rb1 ^G/G, double mutant Rb1 ^G/G ; Cdkn1b ^−/−, and Rb1 ^G/G ; E2f1 ^−/− mice, as well as Rb1 ^G/G ; Cdkn1b ^−/− ; E2f1 ^−/− triple mutant animals. The scale bars represent 20 μm. b Nuclear size from the images in a was determined and the mean size is indicated. Measurements were made from at least 50 nuclei, a, b, c represents statistically different groups as determined by ANOVA followed by Tukey’s test (p < 0.05). c Total number of hepatocytes per 20X field of view was averaged from the indicated genotypes. No statistical differences were observed by AVONA followed by Tukey’s test (p < 0.05) d Nuclei were extracted from livers, stained with propidium iodide, and analyzed for DNA content by flow cytometry. e The relative expression level of six E2F cell cycle target genes from wild type, Rb1 ^G/G, and Rb1 ^G/G ; Cdkn1b ^−/− ; E2f1 ^−/− was determined from RNA extracted from 8-week-old livers. f 8-week-old mice were pulse labeled with BrdU 2 h prior to sacrifice and livers were sectioned and stained for BrdU. The percentage of BrdU positive nuclei was determined. At least 500 nuclei were counted per mouse. All bar graphs represent at least 3 individual experiments, and error bars indicate one standard deviation from the mean. An asterisk represents a statistically significant difference from the wild type control (t test, P < 0.05)