Abstract: FR-PO741
A Human Organoid-on-a-Chip Model Reveals Effects of Flow on Polycystic Kidney Disease Cystogenesis
Session Information
- Cystic Kidney Diseases: Clinical/Translational
November 08, 2019 | Location: Exhibit Hall, Walter E. Washington Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Genetic Diseases of the Kidneys
- 1001 Genetic Diseases of the Kidneys: Cystic
Authors
- Li, Sienna, University of Washington, Seattle, Washington, United States
- Gomez, Ivan G., University of Washington, Seattle, Washington, United States
- Cruz, Nelly M., University of Washington, Seattle, Washington, United States
- Himmelfarb, Jonathan, Kidney Research Institute, Seattle, Washington, United States
- Freedman, Benjamin S., University of Washington, Seattle, Washington, United States
Background
Polycystic kidney disease (PKD) is caused by the loss of polycystin proteins that are posited to act as flow-sensitive mechanosensors on primary cilia, but the effects of flow on cystogenesis are difficult to study in vivo. Human kidney organoids differentiated from pluripotent stem cells provide a genetically specific model of PKD cystogenesis in vitro. We hypothesized that adding flow to this model would exacerbate cyst growth resulting from PKD mutations.
Methods
Kidney organoids were derived from stem cells with bi-allelic, truncating mutations in PKD1 or PKD2, and subjected to fluid shear stress at physiological levels (0.2 dynes/cm2) in microfluidic chips. A static culture condition in the same chips that allowed for simple diffusion of nutrients was used to control for the effects of flow. Fluorescence-labeled glucose was introduced and monitored in time-lapse videos over a 12 hour period, and absorption was quantified using image intensity analysis. Cyst polarity was determined by immunofluorescence for cilia and tight junctions. Cyst expansion was quantified in the presence or absence of transporter modulators such as SGLT2 inhibitors and mannitol.
Results
Cysts in PKD organoids were highly dynamic and increased in size rapidly under flow, compared to static controls. Flow could also be substituted by a diffusive static condition that exposed organoids to an equivalent volume of culture media to achieve a similar effect. PKD cysts in organoids polarized with the apical ciliated surface facing outwards towards the media and exposed to flow, modeling the interior of a tubule in vivo. Expansion of PKD cysts was coupled to glucose transport into epithelial structures and their lumens, while inhibition of glucose transport decreased cyst expansion in a dose-dependent manner.
Conclusion
Our findings suggest that flow, volume, and glucose transport are positive regulators of cyst expansion. Cystogenesis can be enhanced through mechanisms of tubular reabsorption in addition to secretion, and glucose transporters may be crucial to this effect. Collectively, our data support a role for polycystins in regulating the delicate balance of renal transporter function. Therapeutics that reduce flow through the kidney may therefore be beneficial in reducing cyst growth in PKD.
Funding
- NIDDK Support