Abstract
Helium ion scanning microscopy (HIM) is a novel technology that directly visualizes the cell surface ultrastructure without surface coating. Despite its very high resolution, it has not been applied extensively to study biological or pathology samples. Here we report the application of this powerful technology to examine the three-dimensional ultrastructural characteristics of proteinuric glomerulopathy in mice with CD2-associated protein (CD2AP) deficiency. HIM revealed the serial alteration of glomerular features including effacement and disorganization of the slit diaphragm, followed by foot process disappearance, flattening and fusion of major processes, and eventual transformation into a podocyte sheet as the disease progressed. The number and size of the filtration slit pores decreased. Strikingly, numerous “bleb” shaped microprojections were observed extending from podocyte processes and cell body, indicating significant membrane dynamics accompanying CD2AP deficiency. Visualizing the glomerular endothelium and podocyte-endothelium interface revealed the presence of endothelial damage, and disrupted podocyte and endothelial integrity in 6 week-old Cd2ap-KO mice. We used the HIM technology to investigate at nanometer scale resolution the ultrastructural alterations of the glomerular filtration apparatus in mice lacking the critical slit diaphragm-associated protein CD2AP, highlighting the great potential of HIM to provide new insights into the biology and (patho)physiology of glomerular diseases.
Highlights
Scanning electron microscopy (SEM) has been commonly used to examine the complex, three-dimensional structure of the glomerular filtration barrier in the normal kidney and under various pathological conditions in experimental animals and in patients with various glomerular diseases[8,9,10,11]
This method provides a better resolution for imaging glomerular structures. It revealed a novel ridge-like prominence connecting the foot processes which has not been observed by conventional scanning electron microscopy (SEM) or any other electron microscopy techniques before[15]
We report the application of a novel scanning microscopy technology, the Helium Ion microscopy (HIM) in proteinuric glomerulopathy due to the deficiency of a SD protein, CD2-associated protein (CD2AP)
Summary
Kenji Tsuji[1], Teodor G. This method provides a better resolution for imaging glomerular structures It revealed a novel ridge-like prominence connecting the foot processes which has not been observed by conventional SEM or any other electron microscopy techniques before[15]. Our data revealed multiple ultrastructural abnormalities of the glomeruli in Cd2ap-KO mice, including the appearance of numerous “bleb” shaped microprojections, intercellular junctional structure in effaced foot processes and disrupted glomerular endothelium. A complex interface consisting of podocyte, endothelium and glomerular basement membrane was readily visualized by HIM These exciting findings support the utility of this high power scanning microscopy technology in the comprehensive research of glomerular diseases
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