In-Vitro 3D Glomerular System for High-Throughput Biomarker Screening



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Glomerular injury and immune cell infiltration are underlying features involved in multiple renal diseases. Unbiased screens of patient urine and serum have identified a number of potential biomarkers involved in the disease pathogenesis. However, the validation of these biomarkers has been challenging, due to the complicated and time-consuming nature of in-vivo mouse experiments and the inability of 2D culture systems to accurately mimic the in-vivo microenvironments which exist in the renal tissue. We are developing a 3D model of the glomerulus using the renal cells (podocytes, endothelial cells, mesangial cells, and tubule cells) cultured from mouse glomeruli. When these individually cultured cells are brought together in the 3D co-culture, the cell-cell signaling pathways and feedback circuits which exist within these different cells are restored, mimicking the in-vivo environment. The levels of the biomarkers in these cultured cells can be modulated using a lentivirus-mediated gene transduction for overexpression of these genes. Flow cytometry is used to identify the changes in these different cell populations as well as their survival/proliferation. Bone-marrow derived macrophages are also cultured and labeled with fluorescent dyes for modeling immune cell infiltration. The glomerular integrity is measured by quantifying the extent of macrophage infiltration through the endothelial cell/podocyte layer using a confocal microscope. This high-throughput 3D glomerular screening platform will help in identifying the key biomarkers and their relevant signaling pathways involved in disease pathogenesis of multiple renal diseases, and therapeutic interventions targeting these biomarkers could be beneficial in the treatment of these diseases.