Human kidney model
Current 2D cell culture and animal models are insufficient to correctly predict drug effects on human kidney function. While 2D culture systems fail to differentiate and express transporters at physiological levels, there is a major translation gap between animal models and humans. At Mimetas, we have developed perfused epithelial kidney tubules in OrganoPlates®. To this end, we seed renal proximal tubule cells against an extracellular matrix (ECM) gel. Upon initiation of a perfusion flow, a tubular structure is formed. These kidney tubules can be derived from a variety of cell sources and are fully differentiated.
Renal proximal tubule epithelial cells (RPTEC, MDCK or LLC-PK1) are grown against an ECM gel in a three-channel OrganoPlate®, providing access to both the apical and basal side (Fig 1A). The tubules stain positive for ZO-1 (tight junctions) and acetylated tubulin (polarization marker). Cilia point in the direction of the lumen (Fig. 1B) and are absent on the basal, which also demonstrates the correct tissue polarization. The leak-tightness of the boundary is assessed by diffusion of a FITC-dextran dye added to the lumen of the tubule. Boundaries of the PTECs cells form leak-tight barriers and can be maintained for several days for prolonged exposure studies (Fig 1C).
Figure: A) 3-channel layout of the Mimetas OrganoPlate® B) cilia pointing in the direction of the lumen (red) C) Assessment of the leak tightness of proximal tubule cells over time: tubules are leak tight for more than 20 hours.
The renal proximal tubule is responsible for the excretion of a range of compounds, including xenobiotics, and for the reabsorption of valuable compounds from the glomerular filtrate, such as glucose. Various influx and efflux transporters are mediate these functions and are expressed on the basolateral or apical side of the tubule cells. Mimetas has developed a range of fluorescence-based assays to study the activity of specific transporters.
Healthy tubes in an OrganoPlate® are fully leak-tight and may lose their barrier integrity upon compound exposure or disease induction. Barrier integrity of the tubes is assessed by apical perfusion of a dye. Upon disruption of the barrier, e.g. via drug-induced toxicity, the dye will leak towards the basal side, yielding a higher signal in the ECM compartment. This is monitored in real-time and can be combined with other assays to further investigate the mode of toxicity.
The 3D proximal tubules cultured in the OrganoPlate® enable any-throughput toxicity screening and transport studies by real-time imaging. Furthermore, we can establish complex co-culture models to recreate an in vivo-like microenvironment, kidney disease models and study renal clearance.