Integrated Spatial and Single Cellular Profiling in Human IgAN Kidney Biopsies Reveals Novel Cellular Cross-Talk and Stratified Gene Modules of Disease

Background: IgA nephropathy (IgAN) is the most prevalent primary glomerulonephritis worldwide, resulting from glomerular deposition of galactose-deficient IgA immune complexes. Conventional histological scoring has limited prognostic accuracy and prediction of treatment response. To improve disease classification and identify new therapeutics we need better understanding of molecular disease pathways. Integrated glomerular spatial and single nuclei transcriptomic profiling of human renal biopsies may reveal cellular changes and interactions between immune infiltrates and glomerular cells. Methods: Renal biopsy cores from IgAN and pre-implantation living donor patients were processed for single-nuclei RNA sequencing (snRNA-Seq). SnRNA-Seq libraries were integrated and analysed using R software to assess cell composition, cellular communication and differentially expressed genes. Paraffin-embedded renal biopsy sections from 16 patients were profiled via GeoMx for glomerular whole transcriptome, with subsequent expression analysis using R packages and custom scripts. Results: In complementary analysis, snRNA-Seq permits deconvolution of GeoMx glomerular profiles, while GeoMx permits spatial location of snRNA cell populations. Expansion of glomerular immune populations in IgAN is associated with podocyte cell cycling, mesangial Wnt signaling, matrix expansion, and immune activation. Heterogenous glomerulus inflammatory and fibrotic patterns across 166 single glomeruli are linked to gene modules, suggesting disease trajectories. Spatial observations support snRNAseq predicted cell-cell communication between podocyte, mesangial, and immune cell subsets containing T and B cells in the IgAN samples highlighting disease associated immune-glomerular crosstalk in IgAN. Conclusion: Combined single cell and glomerular spatial profiling reveals novel cell specific disease signatures from patient biopsies and highlights cellular crosstalk, and both inter- and intra-sample heterogeneity. We highlight glomerular stratification, with variable immune microenvironment contributing to cellular signaling within diseased glomeruli. This approach is scalable, and results will be integrated to identify novel therapeutic targets and improve disease stratification and classification. Funding: Commercial Support - Novo Nordisk Fellowship Funding, Government Support – Non-U.S.