c-Kit receptor in intercalated cells is required for normal cellular composition of the mature collecting duct

The kidney collecting duct is made up of principal cells (PC), which reabsorb water via apical aquaporin-2 (AQP2), and intercalated cells (IC), which secrete or reabsorb H + via apical (A-IC) or basolateral (B-IC) V-ATPase, thus maintaining water and acid-base homeostasis. The collecting duct is highly plastic even in the adult kidney and the epithelial layer can be altered in response to a variety of pathophysiological cues. For example, during kidney development AQP2-positive cells differentiate into both PC and IC and in the mature collecting duct potassium depletion or treatment with lithium promotes PC differentiation into IC. This decrease in the PC to IC ratio (PC:IC) hinders water reabsorption, and ultimately, leads to the development of nephrogenic diabetes insipidus (NDI). However, the precise mechanism by which epithelial cell fate is determined in the adult kidney collecting duct is not known. The c-Kit signaling axis is required for proliferation, survival, and differentiation of a wide variety of cell types. Although c-Kit and its ligand, Stem Cell Factor/Kit Ligand (KL), can be expressed in the same cell, KL is often expressed in a complementary fashion to c-Kit and signaling occurs between adjacent cells. In the adult kidney collecting duct, c-Kit is highly differentially expressed in A-IC, while its ligand, KL, appears to be most highly expressed in PC by RNA sequencing. However, expression of KL at the protein level has not been examined in the collecting duct. Here, using confocal microscopy we confirmed that in wildtype mice c-Kit is specifically expressed on the basolateral membrane of A-IC, whereas we showed that its ligand, KL, is expressed at the protein level in both PC and A-IC. This finding led us to hypothesize that c-Kit receptor signaling in A-IC is required to maintain the cellular composition of the mature collecting duct. Here, we showed for the first time that c-Kit W sh /W sh mice, which express a mutation in a transcriptional regulatory element upstream of c-Kit, had a significant decrease in c-Kit expression in total kidney lysates by western blotting and PCR. Furthermore, confocal imaging revealed that c-Kit is absent from the basolateral membrane of A-IC from W sh /W sh mice. Thus, W sh /W sh mice are a good loss-of-function model for examining the role of c-Kit in the kidney collecting duct, although they have not previously been used for this. To illuminate the role of c-Kit in the collecting duct we first measured urine pH, which is an indirect measure of proton secretion by A-IC. We found that male W sh /W sh mice presented with a significantly lower urine pH than wildtype mice. We did not observe any gross morphological abnormalities in the collecting duct of these mice by confocal imaging, but c-Kit is known to play a role in cell fate determination. Therefore, we examined the composition of the collecting duct epithelia by quantifying the relative numbers of AQP2-positive PC and B1 V-ATPase-positive IC in the inner medullary collecting ducts via confocal imaging. Intriguingly, male W sh /W sh mice had a lower PC:IC ratio under baseline conditions, which indicates that they have fewer water-absorbing PC and more proton-secreting A-IC, which could explain the abnormally low urine pH we observed in these mice. Taken together, these data indicate that c-Kit expressed in A-IC is involved in the regulation of kidney collecting duct epithelial cellular composition. Our working hypothesis is that c-kit, in conjunction with other signaling molecules, determines the ratio of PC:IC in the mature collecting duct, thus determining the kidneys ability to maintain proper extracellular volume and acid-base parameters. NIH/NIDDK R01DK121848 to D.B. This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.