Figure 3.

Analysis of cellular organization, cell behavior, and cell death at the site of epithelial interconnection in the mammalian kidney. (A1–C2) Anti-laminin immunostaining shows that the basal lamina is discontinuous at the RV to ureteric epithelial interface at RV (A1 and A2) and early S-shaped body (B1 and B2) stages. Pan-cytokeratin labels the collecting duct epithelium (CDE), whereas anti-GFP labels the cap mesenchyme (CM) and RV derivatives (RVd). At the early S-shaped body stage (C1 and C2), a distal cell invasion of an early S-body–derived cell is observed into the ureteric epithelium in an area displaying a punctate laminin distribution (arrow). (D1 and D2) Anticaspase-3 (D1) and TUNEL (D2) labeling fails to detect cell apoptosis at the S-shaped body/ureter junction although apoptotic figures are observed elsewhere (Supplemental Figure 3). (E1–F2) Occludin labeling of tight junctional complexes supports ZO-1 and Par3 analysis of a patent luminal connection between nephron and ureteric epithelium at late S-shaped body stages. (E2) Arrow indicates distal-most cells of RV derivative do not express occludin, whereas the neighboring RV derivative cells express occludin (arrowhead). (F2) Arrow indicates continuous occludin labeling between two epithelial networks. (G1–I2) The absence of phospho-β-catenin (G1–H2) and ZO-1 (I1 and I2), markers of adherens and tight junctional complexes, respectively, highlights the absence of an epithelial organization to the most distal cells of early and mid S-shaped body stages where they interface with the ureteric epithelium (white arrow). Proximal cells of RV derivative have more mature epithelial junctions (white arrowhead). (J1 and J2) Invading distal cells (arrow) do not express Par3.