Hyperactivation of Ha-ras oncogene, but not Ink4a/Arf deficiency, triggers bladder tumorigenesis
J. Clin. Invest. Lan Mo, et al. 117:314 doi:10.1172/JCI30062 [Go to this article.]

Figure 3
The effect of gene dosage of activated Ha-ras on urothelial tumorigenesis. (A) Genotyping by Southern blotting of mice derived from intercrossing of heterozygous mice, showing wild-type mice with the endogenous UPII fragment (lanes 4 and 6), heterozygous mice with Tg/EG ratio of about 1:2 (lanes 1–3, 5, and 8), and homozygous mice with Tg/EG ratio of about 1:1 (lanes 7 and 9). (B) Real-time RT-PCR quantification of activated Ha-ras expression in urothelia of Tg mice, showing that homozygous (+/+) mice expressed nearly twice as much as heterozygous (+/–) mice. The expression level was calculated as ratio in reference to a simultaneously amplified internal GAPDH control. n, number of animals assayed. Bars denote SD. (C) Quantification by real-time PCR of the expression of the transgene-encoded rabbit Ha-ras and the endogenous mouse Ha-ras in heterozygous mice. Note that the levels of expression were roughly equal. (D) Western blot analysis of ras proteins of the wild-type mice (lanes 1 and 2) and heterozygous (lanes 3 and 4) and homozygous (lanes 5 and 6) UPII/Ha-ras-M–transgenic mice. (E) Western blot analysis of p16Ink4a and p19Arf expression in age-matched (6 months) wild-type (lane 1), heterozygous (lanes 2 and 3), and homozygous (lanes 4 and 5) UPII/Ha-ras-M–transgenic mice. Urothelial proteins from UPII/SV40T-transgenic mice were used as positive control (lane 6). (F) Comparison of the tumor-free rate in hetero- or homozygous mice. (G) Morphology of bladder tumors from heterozygotes and homozygotes. Original magnification, top row, ×50; bottom row, ×200.