Human Pluripotent Stem Cell–Derived Kidney Model for Nephrotoxicity Studies

Abstract

Current in vitro models for identifying nephrotoxins are poorly predictive. We differentiated human pluripotent stem cells (hPSCs) into three-dimensional, multicellular structures containing proximal tubule cells (PTCs) and podocytes and evaluated them as a platform for predicting nephrotoxicity. The PTCs showed megalin-dependent, cubilin-mediated endocytosis of fluorescently labeled dextran and active gamma-glutamyl transpeptidase enzymes. Transporters from both the ATP-binding cassette (ABC) and the solute carrier (SLC) families were present at physiological levels in the differentiated cells, but important renal transporters such as organic anion transporter 1 (OAT1), OAT3, and organic cation transporter 2 (OCT2) were present only at lower levels. Radioactive uptake studies confirmed the functional activity of organic cation transporter, novel, type 2 (OCTN2), organic anion transporter polypeptide 4C1 (OATP4C1), and OCTs/multidrug and toxin extrusion proteins (MATEs). When treated with 10 pharmacologic agents as a test of the platform, the known nephrotoxic compounds were distinguished from the more benign compounds by an increase in tubular (PTC, kidney injury molecule 1 (KIM-1), and heme oxygenase 1 (HO-1)) and glomerular (nephrin [NPHS1]/Wilms tumor protein [WT1]) markers associated with nephrotoxicity, and we were able to distinguish the type of nephrotoxin by examining the relative levels of these markers. Given the functions demonstrated and with improved expression of key renal transporters, this hPSC-derived in vitro kidney model shows promise as a platform for detection of mechanistically different nephrotoxins.

Generation of Induced Nephron Progenitor-like Cells from Human Urine-Derived Cells

“…These cells expressed the proximal tubular cell-specific markers CD13, AQP1, and LTL and epithelial maker E-CADHERIN, as determined by qRT-PCR and immunostaining (Figure 5b,c). Moreover, the differentiated cells showed the functional activity of the proximal tubule via uptake of fluorescently labeled dextran as previously reported [40]. While a substantial amount of dextran was accumulated in iNPC-and ESC-differentiated tubular cells, very little dextran was found in the cells induced from UCs (Figure 5d and Figure S6e).…”

Drug-Induced Nephrotoxicity Assessment in 3D Cellular Models

“…Gentamicin [17][18][19][20] Citrinin [17,21] Cisplatin [17][18][19][22][23][24][25][26][27][28][29] Rifampicin [17,30] Acetone [25,28,31] Aspirin [24,27,32] Penicillin G [24,27,33] Tenofovir [26,34] Cyclosporin A [26,35,36] Adriamycin [19,25,37] 4-aminophenol (PAP) [25,38] Colchicine [25,39] Cadmium chloride [40,41] Brush border membrane of the proximal tubules S2 proximal tubular segment Basolateral membrane of proximal tubules Apical membrane of renal proximal tubules S1 and S2 proximal tubular segment Loop of Henle Brush border membrane of the proximal tubules Basolateral mem-brane of proximal tubules Brush border membrane of the proximal tubules/Thick ascending limb of the loop of Henle Brush border membrane of the proximal tubules Loop of Henle S3 proximal tubular segment S1 proximal tubular segment Doxorubicin [17,42,…”

“…The model can correctly identify four renal tubular toxins (gentamicin, citrinin, cisplatin, and rifampicin), as evidenced by the increase in the renal tubular biomarkers KIM-1 and HO-1. When differentiated cells were treated with doxorubicin and puromycin, mainly glomerular toxins, increased levels of NPHS1/WT1 were observed [17]. Kumar et al described a modified suspension culture method for the generation of kidney microorganoids from h-iPSC.…”

How to assess kidney function in oncology patients

“…These include kidneyon-a-chip systems, intravital microscopy of nephrons, 3dimensional organoid cultures, and stem cell-derived nephron models. 51,53,[55][56][57] For example, intravital microscopy has the potential to directly visualize structural and functional changes (including changes in flow, transporter function, and gene/protein expression) in the nephron in living animals over time after the administration of nephrotoxic agents. 46,47 Other methods such as the kidney-on-a-chip systems incorporate both microfluidics, to model flow through the nephron, and 3-dimensional organid cell culture.…”