Fig. 8.

Ablation of Foxd1 cells using diphtheria toxin chain A (DTA) results in significant renal abnormalities. A: gross morphology of kidneys from mice expressing DTA upon Foxd1-cre-mediated recombination (Foxd1-DTA) reveals midline fusion, a failure to separate from the retroperitoneum, and blood vessels visible on the periphery of the kidney cortex. B and C: immunohistochemistry for α-SMA (brown) in control and Foxd1-DTA kidneys. B: at E15.5, control kidneys have well-defined SMC layers surrounding major arteries (arrows) in the interior of the kidney. By contrast, Foxd1-DTA kidneys display areas of diffuse and poorly organized staining within the kidney (arrowheads). However, large, fully coated vessels can be observed along the periphery of the kidney (arrows). At this age, control kidneys also display all of the stages of nephrogenesis, up to and including glomeruli (circles). Meanwhile, Foxd1-DTA kidneys lack mature glomeruli or any of the earlier stages of nephrogenesis, such as comma-shaped or s-shaped bodies. Scale bars: 100 μm C: at E18.5, control kidneys have multiple α-SMA-positive arterioles (arrows) throughout the kidney cortex. Meanwhile, Foxd1-DTA animals have comparatively fewer arterioles. In addition, a large subset of arteries appears to originate from the periphery of the kidney (arrowheads). In contrast to E15.5, Foxd1-DTA kidneys now possess all of the different stages of nephron development, but we note that the nephrogenic zone (brackets) is broader in Foxd1-DTA animals. Scale bars: 50 μm. D: quantification of the orientation of arteries seen in E18.5 Foxd1-DTA kidney demonstrates that nearly two-thirds of vessels observed lay directly underneath or emerge from the capsule. E: average thickness of the smooth muscle layer of renal arteries and arterioles is reduced in Foxd1-DTA animals at E18.5. F: E18.5 Foxd1-DTA kidneys have relatively fewer comma-shaped and s-shaped bodies. G: lineage tracing of Foxd1 cells upon ablation. Foxd1-DTA mice were crossed with mice carrying the R26R allele, which expresses β-galactosidase upon cre-mediated recombination, labeling Foxd1-expressing cells and their derivatives. In control animals at E18.5, the Foxd1-lineage marker labels arterial SMCs and mesangial and interstitial cells. The same cell types are labeled in Foxd1-DTA animals. However, a decrease in the thickness of the Foxd1-lineage SMC layer (arrows) and a marked decrease in the complement of mesangial cells is observed. We also note increased ectopic expression of the lineage marker within tubules (arrowheads). Scale bars: 50 μm. H–I: double staining of Foxd1-DTA kidneys, with LacZ reaction (blue) for Foxd1-lineage cells and immunohistochemistry (brown) for α-SMA. As seen in this afferent arteriole extending from the glomerulus, a subset of arteriolar SMCs is positive for both the lineage marker and α-SMA (black arrows), while other SMCs are positive only for α-SMA and not the lineage marker (white arrows). I: some mesangial cells are double-positive for both markers (black arrows), while others are only positive for α-SMA but negative for the lineage marker (white arrows). Scale bars: 50 μm.