Super-Resolved Three-Dimensional (3D) Imaging and Analysis of Whole Mouse Glomeruli via GloMAP

Background: Glomerular structure is central to kidney function, yet traditional imaging relies on thin sections or modest spatial resolution, limiting the ability to study whole glomeruli as 3D functional units. To address this challenge, we developed GloMAP, a pipeline combining super-resolution optical microscopy and human-assisted computational segmentation to reconstruct whole mouse glomeruli at 100nm resolution. Methods: We used a novel fluorescent tricolor stain to label tissue sections and we imaged whole hydrogel-expanded glomeruli by confocal microscopy. Manual and machine-learning-assisted segmentation defined six structural compartments (e.g., GBM, Bowman’s capsule) and enabled classification of cell types. These data were used to analyze volume fractions, surface areas, basement membrane thicknesses, and other properties. Results: 24 glomeruli from young adult, aged, and experimental FSGS mice were reconstructed, allowing analysis of glomerular compartments and structures and classification of the 4 principal glomerular cell types with >90% accuracy. This revealed age- and disease-associated changes, including segmental basement membrane thickening, altered mesangial structure, and altered cell type proportions. Global maps of GBM and BCBM thickness revealed distinct remodeling patterns across conditions. In experimental FSGS glomeruli, we observed multilayered BCBM, podocyte hypertrophy, and parietal epithelial cell migration (Fig. 1). Conclusion: GloMAP enables high-resolution, 3D reconstruction of whole glomeruli using accessible methods. It is a powerful platform for studying whole glomerular architecture and it provides a foundation for integrative modeling and therapeutic evaluation. Funding: NIDDK Support - NIDDK Support, NIDDK SupportFig. 1. Representative segmental pathological patterns from an experimental FSGS mouse glomerulus.