Analysis of early nephron patterning reveals a role for distal RV proliferation in fusion to the ureteric tip via a cap mesenchyme-derived connecting segment

Abstract

While nephron formation is known to be initiated by a mesenchyme-to-epithelial transition of the cap mesenchyme to form a renal vesicle (RV), the subsequent patterning of the nephron and fusion with the ureteric component of the kidney to form a patent contiguous uriniferous tubule has not been fully characterized. Using dual section in situ hybridization (SISH)/immunohistochemistry (IHC) we have revealed distinct distal/proximal patterning of Notch, BMP and Wnt pathway components within the RV stage nephron. Quantitation of mitoses and Cyclin D1 expression indicated that cell proliferation was higher in the distal RV, reflecting the differential developmental programs of the proximal and distal populations. A small number of RV genes were also expressed in the early connecting segment of the nephron. Dual ISH/IHC combined with serial section immunofluorescence and 3D reconstruction revealed that fusion occurs between the late RV and adjacent ureteric tip via a process that involves loss of the intervening ureteric epithelial basement membrane and insertion of cells expressing RV markers into the ureteric tip. Using Six2-eGFPCre x R26R-lacZ mice, we demonstrate that these cells are derived from the cap mesenchyme and not the ureteric epithelium. Hence, both nephron patterning and patency are evident at the late renal vesicle stage.

Wilms Tumor Suppressor, WT1, Suppresses Epigenetic Silencing of the β-Catenin Gene

“…As a prelude to nephrogenesis, WT1 and EZH2 expression is co-expressed in progenitor cells of the cap mesenchyme, but only WT1 is sustained in the S-shaped body (21,22,30). This suggests either that the effect of WT1 on EZH2 transcript level is delayed or that additional factors contribute to the full downregulation of EZH2 (21,22).…”

Renal organogenesis

“…Lineage tracing verified the cap mesenchyme as the origin of the cells forming this connection segment. 21 Coincident lumens were seen shortly after this time, indicating that this is an early event in nephron differentiation.…”

“…21 Each nephron must fuse with the ureteric epithelium to form a patent lumen, allowing urinary filtrate to exit the kidney. Little was known about this process.…”

“…Key members of the Bmp (Bmp2), Notch (Jag1, Dll1) and Wnt (Dkk1, Wnt4) signaling pathways displayed differential distal RV expression. 21 This portion of the RV also showed differential expression of cyclin D1 (Ccnd1), a marker of the G 1 phase of the cell cycle. Based on this observation, quantification of relative proliferation between distal and proximal RV was performed using the mitotic marker pHH3.…”

“…20 Here, our approach was to identify a set of genes with broader spatial expression domains but whose expression was highest in the renal vesicle. 21 Detailed section in situ …”