Transformative Tools for Imaging Tissue Ultrastructure in Diabetic Kidney Disease
This proposal seeks to apply and adapt the next-generation super-resolution optical microscopy method expansion microscopy (ExM) to the study of human diabetic kidney disease, the most common cause of end-stage renal disease in the U.S. In ExM, fluorescent labels on fixed specimens are linked to a swellable polymer hydrogel that is physically expanded with remarkably low distortion. The process allows features closer than the diffraction limit of light (~250 nm) to become resolvable in the expanded specimen and achieves ~65 nm spatial resolution on thick or thin specimens with ordinary confocal microscopes. This greatly improved spatial resolution is sufficient to reveal key ultrastructural features of the kidney including glomerular foot process width and glomerular basement membrane width while also providing molecular details such as extracellular membrane composition or presence of immune deposits, etc. Importantly, ExM makes use of standard optical microscopes that are already familiar and widely available, together with inexpensive sample processing methods, so that the method has the potential to become widely used in research and clinical settings for the study or diagnosis of kidney diseases, including DKD. Our specific aims are: 1) to develop and optimize procedures to study fresh kidney specimens using ExM; 2) to adapt these procedures to study archived paraffin-embedded tissue by ExM; and 3) to perform correlative ExM and electron microscopy imaging as a robust validation of our methodology.

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