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. 2006 Feb 13;172(4):529–540. doi: 10.1083/jcb.200507081

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Copyright © 2006, The Rockefeller University Press

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Figure 6. — Early onset of cellular senescence in Bub3/Rae1 double haploinsufficient MEFs. (A) Analysis of the percentages of cell death in Bub3^+/−/Rae1^+/− and wild-type MEFs cultures by TUNEL assay (n = 3 MEF lines per genotype). (B) Percentages of Bub3^+/−/Rae1^+/− and wild-type MEFs that are in the early stages of apoptosis (annexin V⁺/PI⁻ cells), in the late stages of apoptosis, already dead (annexin V⁺/PI⁺ cells), that undergo nonapoptotic cell death (annexin V⁻/PI⁺ cells), or that are dead (PI⁺ cells). We used 3 MEF lines per genotype. We note that the differences between Bub3^+/−/Rae1^+/− and wild-type MEFs were not statistically significant using an unpaired t test. (C and D) Growth curves of wild-type, Bub3^+/−, Rae1^+/−, and Bub3^+/−/Rae1^+/− MEF cells at P3 (C) and P7 (D). 1.5 × 10⁵ cells of indicated passages were seeded in duplicate on day 0 and counted for five consecutive days. Lines represent three independent MEF lines. Note that Bub3^+/−/Rae1^+/− MEF cells have reduced proliferation potential at P7, whereas single heterozygous Bub3^+/− and Rae1^+/− MEFs do not. (E) Percentages of wild-type, Bub3^+/−, Rae1^+/−, BubR1^H/H, and Bub3^+/−/Rae1^+/− MEF cells that were positive for SA β-galactosidase activity at the indicated passages (n = 3 lines for each genotype at each passage). Asterisks in D and E mark values that are significantly different from those of wild-type mice using an unpaired t test. Error bars represent SD.