Student McMaster University Waterdown, Ontario, Canada
Abstract Body : Introduction: Nephrons are the functional units of the kidney and are essential for homeostasis. During early nephron formation, nephron progenitor cells migrate and tightly cluster together to form renal aggregates. These aggregates will undergo significant changes in cell morphology, develop cell membrane specializations (i.e., cellular projections), undergo mesenchymal-to-epithelial transition to form a renal vesicle, and eventually mature into a functional nephron. The morphological changes occurring during kidney development rely on the carefully coordinated regulation of actomyosin dynamics. Shroom3 modifies epithelial cell shape via the regulation of actomyosin dynamics and is strongly associated with kidney disease. In embryonic humans and mice, Shroom3 has the highest level of expression in the nephron progenitor cells. Shroom3 null mice exhibit marked defects in maturing nephrons and exhibit reduced nephron numbers, however, how Shroom3 contributes to abnormal nephron formation is unknown.
Methods and Results: Hematoxylin and Eosin staining and Six2 immunofluorescence of embryonic day (E)13.5 kidneys from Shroom3 null mice revealed abnormal nephron progenitor clustering, renal aggregate formation, and a 2-fold reduction in nephron progenitor number. The nephron progenitor cells were loosely clustered with a gap between the ureteric epithelium and nephron progenitors that was 3-fold greater than the wild type. In the wild type, cellular projections emanated from the leading edge of the nephron progenitor cells towards the ureteric epithelium as shown by GM130 and NCAM immunofluorescence. In contrast, the Shroom3 null kidneys exhibited a random orientation of the leading edge and very few cellular projections were observed. Transmission electron microscopy of wild type mice identified the cellular projections as filopodia and lamellipodia. In contrast, the Shroom3 null mice demonstrated poorly formed or absent filopodia and lamellipodia. The Shroom3 null mice also exhibited poor epithelial organization of renal vesicles that often exhibited abnormal epithelial polarization.
Conclusion and Significance: These studies demonstrate Shroom3’s essential role in regulating the nephron progenitor cell morphology, and cell projection formation. As these projections are essential for cellular communication, our studies suggest that a loss of Shroom3 disrupts cellular communication during kidney development. Shroom3 is the second most significant gene associated with kidney disease in humans and our results provide evidence that adult-onset kidney disease may result from abnormal nephron development during the embryonic period, leading to abnormally formed nephrons an/or reductions in nephron number.
