Abstract
The glomerulus exercises its filtration barrier function by establishing a complex filtration apparatus consisting of podocyte foot processes, glomerular basement membrane and endothelial cells. Disruption of any component of the glomerular filtration barrier leads to glomerular dysfunction, frequently manifested as proteinuria. Ultrastructural studies of the glomerulus by transmission electron microscopy (TEM) and conventional scanning electron microscopy (SEM) have been routinely used to identify and classify various glomerular diseases. Here we report the application of newly developed helium ion scanning microscopy (HIM) to examine the glomerulopathy in a Col4a3 mutant/Alport syndrome mouse model. Our study revealed unprecedented details of glomerular abnormalities in Col4a3 mutants including distorted podocyte cell bodies and disorganized primary processes. Strikingly, we observed abundant filamentous microprojections arising from podocyte cell bodies and processes, and presence of unique bridging processes that connect the primary processes and foot processes in Alport mice. Furthermore, we detected an altered glomerular endothelium with disrupted sub-endothelial integrity. More importantly, we were able to clearly visualize the complex, three-dimensional podocyte and endothelial interface by HIM. Our study demonstrates that HIM provides nanometer resolution to uncover and rediscover critical ultrastructural characteristics of the glomerulopathy in Col4a3 mutant mice.
Highlights
Ultrafiltration of plasma through a complex glomerular filtration barrier, consisting of podocytes, endothelial cells and glomerular basement membrane (GBM), represents the most important function of the kidney[1,2]
Makino et al visualized gaps in the GBM and red blood cells passing through these gaps in hematuric animals with a combination of transmission electron microscopy (TEM) and Scanning Electron Microscopy (SEM), uncovering the cause of hematuria associated with kidney disease[24]
Foot process effacement detected by TEM and SEM has become a hallmark of proteinuric glomerulopathy/podocytopathy[25]
Summary
Three-dimensional view of podocytes in Col4a3 mutant mice. 4–5 month old Col4a3−/− (Alport) mice lacking the collagen α3α4α5(IV) network and wild-type (WT) and heterozygous (Col4a3+/−) control mice were used for scanning HIM. The foot processes of WT and Col4a3+/− animals were mostly of uniform size and well organized (Fig. 3a-i,ii) They appeared wider in Col4a3−/− mice (Fig. 3a-iii), which is consistent with our observations by TEM. There was a recent report of “ridge-like prominences” observed by block-face scanning electron microscopy[27] These “ridge-like prominences” project from the basal side of the podocyte cell body and major processes, and are closely attached to the GBM along the foot processes. In the late stage glomeruli in Col4a3−/− mice, HIM revealed that the distinction between primary and foot process morphology as seen in the WT animals (Fig. 5a) was lost and was replaced by broadly effaced podocyte processes that formed large and flattened sheets covering capillaries below. This opens up a timely opportunity to uncover and rediscover anatomic features of various glomerulopathies for disease diagnosis, differentiation and more importantly, for the understanding of the specific cellular and molecular processes associated with sophisticated morphological features of various glomerulopathies
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
