Profiling of Potentially Harmful Circulating Factors in the Serum of Patients with FSGS Using Human Induced Pluripotent Stem Cell (iPSC)-Derived Podocytes

Background: Focal segmental glomerulosclerosis (FSGS) presents significant challenges due to its complexity and variable clinical manifestations. Current evidence suggests that FSGS pathogenesis can be caused by circulating factors (CF), present in the serum of patients. However, the nature and quality of these potential factors is still a topic of intense research. A hallmark and early indicator of podocyte damage in FSGS is the rearrangement of the actin cytoskeleton, which compromises podocyte structure and function, leading to foot process effacement and cell loss. Methods: We used sophisticated differentiation methods to generate homogeneous cultures of human iPSC-derived podocytes. The resulting multi-well plate system recapitulates uniquely the expression of specific mature podocyte markers (Nephrin and Podocin). The cells equally recapitulate several of the morphological and functional properties of their primary counterparts. We took advantage of this new platform to develop a specific readout for the direct profiling of FSGS serum from patients. Results: Serum samples of multiple diseased and healthy individuals were incubated with iPSC-derived podocytes and characterized with respect to their effects on actin and cytoskeleton structure. High-content imaging and script-based analysis were used to quantify structural changes and potential cell death (using apoptosis readouts). The new iPSC-podocyte system successfully discriminated against different patient sera, potentially containing CF detrimental to their structural integrity. One specific sample of FSGS serum was able to significantly modulate podocyte actin architecture and resulted in measurable changes in cytoskeleton organization. Conclusion: Identification of particularly harmful serum can help understanding the pathogenic mechanisms at play in FSGS patients. Further multiomic and functional analyses are warranted to characterize molecularly the traits of these pathogenic drives. For now, the combination of human iPSC derived podocytes and high-content imaging holds promising applications in drug discovery, for kidney transplantation, graft compatibility and FSGS relapses in organ recipients.