Fig. 3.

Analysis of the urothelium. (A) Comparison of bladder urothelium in p63–/– and control littermates (18.5 dpc), showing the presence of a single layer of cells with ample eosinophilic cytoplasm in p63–/– embryos. Similarly to umbrella cells of wild-type mice, these cells express Uroplakin III (UP III). p63-positive basal and intermediate cells of wild-type urothelium are shown in Inset. (Magnification: ×200X). (B) X-gal staining and p63 immunostaining in the bladder of a partially rescued p63–/– chimera (18.5 dpc), showing partial absence of the basal and intermediate cell layers (arrows). In the rescued epithelium, p63-positive basal and intermediate cell layers are exclusively formed by p63+/+ β-gal-positive cells. The umbrella cell layer consists of p63–/– β-gal-negative cells. Lower represents higher magnification (×400) of the marked regions in top panels (×40). (C) Comparison of X-gal staining of the bladder epithelium of p63–/– chimeras, control chimeras, and ROSA26 mice at 7 weeks. Center and Right are higher magnifications (×200) of marked regions in Left (×40). The asterisk indicates the bladder lumen, the black arrowhead indicates basal/intermediate cells, and the open arrowhead indicates umbrella cells. In control chimeras, both the basal/intermediate and the umbrella cell layers are occupied by β-gal-positive and -negative cells. In p63–/– chimeras, only p63+/+ β-gal-positive cells are present in the basal and intermediate cell layers. Approximately 85% of the cells that constitute the umbrella cell layer are p63–/– (β-gal negative). All urothelial cells of ROSA26 mice display β-gal activity. (D) X-gal staining of the urothelium lining the ureters (×200). Similarly to the bladder urothelium, p63–/– β-gal-negative cells are present in the umbrella layer of p63–/– chimeras. In the control chimera, β-gal-positive umbrella cells are observed above β-gal-negative basal and intermediate cells. The ureter of ROSA26 mice is entirely β-gal positive.