The laboratory of James Pullman, MD, PhD, is exploring the use of a new method to diagnose glomerular disease, a complex area of renal pathology that requires more than the routine light microscopy (LM) that suffices for diagnosis in other organs.
Glomerular disease pathology uses electron microscopy (EM) to image structures too small for LM to resolve, as well as immunofluorescence microscopy (IFM) to identify and localize specific proteins. Super Resolution Microscopy (SRM) is a group of new technologies that combines the molecular mapping capabilities of IFM with the resolving power of EM, and can provide information similar to and likely beyond what either can offer.
Dr. Pullman is working with a team of physicists from the University of St. Andrews in Scotland, led by Professor Kishan Dholakia. They have employed Structured Illumination Microscopy (SIM), a type of SRM with the advantage of using specimen preparations similar to those of conventional IFM. Initially, the investigators examined the pathology of the podocyte, one of three cell types in the glomerulus. The podocyte is distinguished by its “foot processes,” unusual cytoplasmic extensions which retract into the cell body in diseases associated with proteinuria, or nephrotic syndrome.
Until now, EM, and not LM or IFM, provided the only way to visualize foot processes. The Pullman Laboratory has shown that SIM, with an immunofluorescent-labeled antibody bound to the protein podocin, imaged the outlines of foot processes in normal and diseased podocytes, and matched the changes seen by EM. In addition, the large field of view provided by SIM, an order of magnitude greater than that of EM, revealed novel changes in foot processes that could not be seen with EM.
Dr. Pullman and his colleagues are further analyzing these changes to understand their significance in nephrotic disease. In addition to podocyte pathology, they’re using SIM to map disease-related changes in other glomerular proteins, cells and extracellular matrices, as well as molecules that infiltrate the glomerulus and cause disease, such as immune complexes.