Invasion of Distal Nephron Precursors Associates with Tubular Interconnection during Nephrogenesis

Abstract

Formation of a functional renal network requires the interconnection of two epithelial tubes: the nephron, which arises from kidney mesenchyme, and the collecting system, which originates from the branching ureteric epithelium. How this connection occurs, however, is incompletely understood. Here, we used high-resolution image analysis in conjunction with genetic labeling of epithelia to visualize and characterize this process. Although the focal absence of basal lamina from renal vesicle stages ensures that both epithelial networks are closely apposed, we found that a patent luminal interconnection is not established until S-shaped body stages. Precursor cells of the distal nephron in the interconnection zone exhibit a characteristic morphology consisting of ill-defined epithelial junctional complexes but without expression of mesenchymal markers such as vimentin and Snai2. Live-cell imaging revealed that before luminal interconnection, distal cells break into the lumen of the collecting duct epithelium, suggesting that an invasive behavior is a key step in the interconnection process. Furthermore, loss of distal cell identity, which we induced by activating the Notch pathway, prevented luminal interconnection. Taken together, these data support a model in which establishing the distal identity of nephron precursor cells closest to the nascent collecting duct epithelium leads to an active cell invasion, which in turn contributes to a patent tubular interconnection between the nephron and collecting duct epithelia.

De novo lumen formation and elongation in the developing nephron: a central role for afadin in apical polarity

“…Interestingly, there does not appear to be direct lumen extension from ureteric bud tips. Ultimately, we find the nephron-ureteric bud lumen becomes continuous at the S-shaped body stage, which is consistent with recently published data (Kao et al, 2012). Our data also show that whereas afadin is required for lumen initiation in renal vesicles, at later stages many developing nephrons devoid of afadin will eventually form numerous small lumens.…”

“…Ultimately, the nephron-UB lumen became continuous at the Sshaped body (SB) stage ( Fig. 2I-IЈ), as has recently been reported by others (Kao et al, 2012). We conducted these experiments using sections fixed with either 4% paraformaldehyde or Nakane (not shown), suggesting these lumen morphologies are unlikely to be an artefact of fixation.…”

The luminal connection

“…16 In the mammalian kidney, two recent studies show that the basal lamina is largely discontinuous at the future site of luminal interconnection between the renal vesicle and late-S shaped nephron precursor stages. 17,18 It is possible that the punctate distribution of the basal lamina may provide an inhibitory cue by an unknown signaling mechanism that may act in preventing luminal interconnection. Since the intervening basal lamina is largely denuded at the future interconnection zone, it is less likely that breakdown of basal lamina alone achieves luminal interconnection in the developing kidney.…”

Growing New Kidneys from Embryonic Cell Suspensions

“…In humans, activating mutations in the CaSR gene (Casr) are associated with hypocalcemia with hypercalciuria 2 and, in certain circumstances, a Bartter-like phenotype, 3 with renal salt wasting, alkalosis, and an increase in plasma renin and aldosterone levels. Conversely, heterozygous inactivating mutations in the Casr lead to familial hypocalciuric hypercalcemia (FHH), a relatively benign and asymptomatic condition.…”

“…2 By embryonic day (E) 10.5 in the mouse, the ureteric bud evaginates from the nephric duct and contacts the metanephric blastema to initiate the process of nephrogenesis. 3,4 Thus, tissue harvesting at E11.5 captures the developmental window when outgrowths of ureteric bud are branching and beginning to induce metanephric mesenchymal progenitor cells to differentiate. Evidence suggests that this midembryonic age of tissue harvesting gives superior results over late time points, probably because of greater tissue plasticity and growth potential as well as lower immunogenicity.…”