PhD Student Istituto di Ricerche Farmacologiche Mario Negri IRCCS Milano, Italy
Abstract: The glomerular filtration barrier (GFB), composed of podocytes, glomerular basement membrane (GBM) and fenestrated endothelial cells, is a highly specialized structure responsible for blood filtration in the kidney. Reconstructing the GFB in vitro has long been a challenge due to its intricate three-dimensional architecture and unique biochemical and mechanical properties. This study aimed to establish an efficient glomerulus-on-a-chip model for studying glomerular physiology and pathology employing human induced pluripotent stem cells (hiPSCs). To this end, we first successfully differentiated hiPSCs into podocytes and glomerular endothelial cells, the cellular components of GFB. These cells were then co-cultured on a coated porous membrane of a transwell integrated into a 3D millifluidic system designed to replicate key properties of the mechanical and biochemical microenvironment of the human glomerulus. To simulate physiological shear stress, and promote cell adhesion, alignment and maturation, controlled flow conditions were optimized by using a peristaltic pump linked to this 3D millifluidic system. RT-qPCR experiments and immunofluorescence analysis confirmed the proper formation of the GFB, assessing the expression of specific markers, including nephrin and podocin for podocytes, VE-cadherin and vWF for endothelial cells, and laminin and collagen for GBM deposition. Finally, we used this platform to model pathological conditions such as nephrotoxic injury and Alport Syndrome using patient-specific hiPSCs. Studies on filtration properties and selective permeability validated the reliability of these systems to mimic in vitro human glomerulus diseases. This glomerulus-on-a-chip represents a significant step forward in the development of human-relevant models for renal research. It provides a versatile tool for investigating disease mechanisms, enabling personalized medicine approaches, and advancing drug discovery, addressing the growing need for alternatives to animal models.
