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The glomerulus is the filtration unit of the kidney. Injury to any component of this specialised structure leads to impaired filtration and eventually fibrosis and chronic kidney disease. Current two and three dimensional (2D and 3D) models that attempt to recreate structure and interplay between glomerular cells are imperfect. Most 2D models are simplistic and unrepresentative, and 3D organoid approaches are currently difficult to reproduce at scale and do not fit well with current industrial drug-screening approaches. Here we report a rapidly generated and highly reproducible 3D co-culture spheroid model (GlomSpheres), better demonstrating the specialised physical and molecular structure of a glomerulus. Cocultured using a magnetic spheroid formation approach, conditionally immortalised (CI) human podocytes and glomerular endothelial cells (GEnCs) deposited mature, organized isoforms of collagen IV and Laminin. We demonstrate a dramatic upregulation of key podocyte (podocin, nephrin and podocalyxin) and GEnC (pecam-1) markers. Electron microscopy revealed podocyte foot process interdigitation and endothelial vessel formation. Incubation with pro-fibrotic agents (TGF-β1, Adriamycin) induced extracellular matrix (ECM) dysregulation and podocyte loss, which were attenuated by the anti-fibrotic agent Nintedanib. Incubation with plasma from patients with kidney disease induced acute podocyte loss and ECM dysregulation relative to patient matched remission plasma, and Nintedanib reduced podocyte loss. Finally, we developed a rapid imaging approach to demonstrate the model’s usefulness in higher throughput pharmaceutical screening. GlomSpheres therefore represent a robust, scalable, replacement for 2D in vitro glomerular disease models.
|Publication status||Published - 2 Dec 2021|
Bibliographical noteFunding Information:
This work was part funded by a BBSRC CASE studentship jointly with UCB pharmaceuticals (Slough). Additional funding was provided by Wellcome Trust ISSF, Medical Research Council (MR/R013942/1), the Elizabeth Blackwell Institute (TRACK) and the Nephrotic Syndrome Trust’. M.C. is funded by the Royal Thai Government Scholarship program. Confocal (BBSRC Alert 13 capital grant BB/L014181/1), Light sheet (Wellcome Trust), TEM and SEM (BBSRC 17ALERT) imaging was performed at the Wolfson Bioimaging Facility. TEM and SEM sample preparation and imaging was performed by Dr. Chris Neal and Mrs Judith Mantell.
© 2021, The Author(s).
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Wolfson Bioimaging Facility
Mark Jepson (Manager)Faculty of Life Sciences