Novel Gravity Driven Microfluidic System Development for Study of the Human Kidney

Introduction:: Chronic kidney disease (CKD) and acute kidney injury (AKI) are growing ailments in populations around the world. In the U.S. alone 1 in 15 Americans is estimated to be living with CKD¹. CKD, while manageable, drastically affects quality of life as it is linked to decreased immune response, heart disease, valve failure, and other terminal illnesses. There are few models that accurately depict kidney function outside of the human body. Microphysiological systems (MPS) provide a method to test new drugs and/or drug combinations, and to better understand disease progression. While few models have shown that this technology can demonstrate the functional components of a human kidney, in work completed by Zhang and Mahler, kidney filtration is demonstrated through the combination of a proximal convoluted tubule and glomerulus on a chip system². This system utilizes a peristaltic pump to facilitate the circulation of the blood and waste streams in way that mimics the human body. The overall goal of this work creating a novel and more efficient method to recreate kidney function for diagnostics and drug testing. This work outlines the fabrication and characterization of a simplified, novel, gravity driven device that can sustain kidney-like cell conditions.

Materials and Methods:: Human umbilical vein endothelial cells (HUVEC) were used as a cell model to determine viability of cells in the newly created gravity-driven MPS. Devices were drawn in Alibre CAD software and manufactured out of a clear, ABS-like resin (BioMed Clear by FormLabs). Key features of this device include a low fluid volume and the ability to generate flow using gravity with a custom spinning platform. The novelty of these devices is their use of gravity to produce flow. Further, gravity driven devices require less space, materials, and technical expertise to operate. The physical devices can be used for a longer period of time, as lower amounts of stresses are applied. Devices were coated with 1.5mL of poly-D-lysine at a stock concentration of 0.1mg/mL for 7 minutes, followed by a fibronectin coating of 1.5mL at a concentration of 5µg/cm² for 1 hour a room temperature. Cells were added at 200µL per device at a concentration of of 50,000 cells/mL. Devices were spun at a 45° angle both continuously and at an interval of 90s and kept in an incubator at 37°C and 5% CO₂. After 7 days of operation, cells were stained for viability or VE Cadherin to probe cell-cell junction formation. Images were taken via Zeiss fluorescence microscope.

Results, Conclusions, and Discussions:: Results and Discussion: Current studies have demonstrated that cells adhere to the printed devices. These cells are viable after a 7 day period within the spinning device, however they are not generating strong cell-cell junctions, as noted in Figure 1A and 1B. The lack of cell-cell junctions suggests while the cells are surviving, they are either seeded at too low of a density or are not spreading. Results from the materials testing of the Biomed Clear resin are still inconclusive (Fig. 1 panel C). New tests are still pending with plans to image for both cell viability and ECM production. Additionally, testing with devices will continue for determining continued viability within the device channels.

Conclusion: Results currently indicate that the new gravity driven MPS devices are capable of supporting cells for 7 days of culture. Future work will focus on incorporation of kidney-specific cells. This device will be useful for demonstrating kidney function on a small scale for drug toxicity testing. Further, continued works with the device to develop a filtration layer will be able to mimic the barrier created in the glomerulus and in the proximal convoluted tubule. With successful incorporation of kidney-specific cells a multistep device to mimic kidney filtration can be conceptualized.

Acknowledgements (Optional): : GFSD/GSME Fellowship, Graduate Fellowships for Stem Diversity & NIST

References (Optional): : [1] Chronic Kidney Disease in the United States, 2021. (2022, July 12). Center for Disease Control and Prevention.

[2] Zhang, S. Y., & Mahler, G. J. (2023). A glomerulus and proximal tubule microphysiological system simulating renal filtration, reabsorption, secretion, and toxicity. In Lab on a Chip (Vol. 23, Issue 2, pp. 272–284). Royal Society of Chemistry (RSC). https://doi.org/10.1039/d2lc00887d