Deletion of Gapvd1 Sensitizes Murine Podocytes to Adriamycin-Induced Injury

Background: DNA variants in GAPVD1, an activator of endosomal protein Rab5, are a monogenic cause of nephrotic syndrome. The encoded protein interacts with nephrin and previous studies in podocyte-like nephrocytes in Drosophila suggested a crucial function for slit diaphragm integrity. However, the role of this gene remained unexplored in mammalian animal models. Methods: To generate a conditional knockout, we established a Gapvd1 allele with loxP sites flanking exon 4. Gapvd1fl/fl mice were crossed with Nphs2-Cre for podocyte-specific deletion and Six2-Cre for deletion in nephron progenitor cells. To apply toxicity stress, sixteen week old Nphs2-Cre; Gapvd1fl/fl mice and control animals (Gapvd1fl/fl ) were treated with intravenous Adriamycin. For phenotypic characterization, we determined serum creatinine and urinary albumin-to-creatinine ratios (ACR) and further applied histochemistry, immunofluorescence and electron microscopy. Results: In situ hybridization indicated a successful conditional knockout of Gapvd1. However, unstressed animals using Nphs2-Cre or Six2-Cre for deletion of Gapvd1 revealed no overt phenotype including changes in glomerular architecture, renal function and urinary ACR. In contrast, Adriamycin-injected Nphs2-Cre, Gapvd1fl/fl mice presented with elevated proteinuria compared to the control group. Periodic acid-Schiff staining of kidney sections indicated focal segmental glomerulosclerosis in stressed knockout animals, which further exhibited significantly less WT1-positive podocytes per glomerulus in immunofluorescence. Analysis of glomeruli with scanning electron microscopy revealed shortened and partially fused foot processes and transmission electron microscopy confirmed foot process effacement. Conclusion: Gapvd1 seems dispensable for podocyte homeostasis in unstressed mice, likely due to compensation by another Rab5-regulating protein. However, loss of Gapvd1 sensitized animals to Adriamycin-induced podocyte injury resulting in podocyte loss and proteinuria. Funding: Government Support – Non-U.S.