Sec10 is necessary for maintenance of the uroplakin barrier in the developing ureter urothelium.

<p>Embryos were collected after timed-matings to evaluate ureters at the UPJ prior to formation of the obstruction, which occurs between E17.5 –E18.5. (A, B) Immunostaining for E-cadherin (green) and SMA (red) at the UPJ in <i>Sec10</i>^<i>FL/FL</i>;<i>Ksp-Cre</i> and control E16.5 ureters confirmed initial patency of the mutant UPJs. (C, D) Immunostaining for uroplakin-3 (red) and vimentin to mark the mesenchymal cells (green) revealed that at E16.5, <i>Sec10</i>^<i>FL/FL</i>;<i>Ksp-Cre</i> ureters already demonstrated patchy uroplakin-3 localization. Areas of urothelium with noticeably absent uroplakin-3 are marked with arrows (D), while in control ureters, uroplakin-3 was distributed all around the luminal surface (C). (E, F) By E17.5, staining for E-cadherin (green) and SMA (red) revealed that the urothelial layer was visibly abnormal in morphology and consisted of mostly a single layer. (G, H) At E17.5, uroplakin-3 was strongly detectable in all control ureters, but completely absent in <i>Sec10</i>^<i>FL/FL</i>;<i>Ksp-Cre</i> ureters around the UPJ. (I, J) To measure proliferation in the smooth muscle layers of developing ureters at the UPJ region, we immunostained for Ki67, which is expressed only in mitotic cells. (K) Proliferation rates were calculated from counting all Ki67⁺;SMA⁺ cells and Ki67^-;SMA⁺ smooth muscle cells in <i>Sec10</i>^<i>FL/FL</i>;<i>Ksp-Cre</i> and control ureters at E16.5 and E17.5. Proliferation rates were compared between mutant and control ureters by student t-tests, with means ± SD and n’s for each group shown. In all images, nuclei were counterstained with DAPI.