Kidney Week

Abstract: SA-PO558

Transport and Barrier Function in Cultured Primary Tubule Cells Are Preserved after Brief Cold Storage

Session Information

• Bioengineering
  October 26, 2024 | Location: Exhibit Hall, Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Bioengineering

• 400 Bioengineering

Authors

• Evans, Rachel C., Vanderbilt University Medical Center, Nashville, Tennessee, United States

Rachel C. Evans

• Roy, Shuvo, University of California San Francisco, San Francisco, California, United States

Shuvo Roy, PhD

• Humes, H. David, University of Michigan Michigan Medicine, Ann Arbor, Michigan, United States

H. David Humes, MD, DrMed

• Fissell, William Henry, Vanderbilt University Medical Center, Nashville, Tennessee, United States

William Henry Fissell, MD

Group or Team Name

• The Kidney Project.

Background

Implantable bioartificial kidneys will need to be distributed to clinical sites distant from the point of manufacture. Renal allografts are typically shipped cool surrounded by ice-water baths with or without perfusion, while suspended cells are typically shipped frozen in dry ice. Freezing a device to -78^oC is likely impractical as it subjects a device to destructive mechanical stresses from thermal expansion, and there may be incomplete survival of cells upon thawing, jeopardizing function. We simulated cold transport by examining survival and function of primary renal proximal tubule cells in monolayer culture after storage at 5^oC.

Methods

Primary human renal proximal tubule cells were grown on permeable supports in serum-supplemented 50:50 DMEM:F12 with an inhibitor of TGF-βR1 on a rotary shaker table at 37^oC and 5%CO₂/air for 28 days. Baseline apicobasal barrier was assessed by leak of TRITC-labeled dextrans from apical to basolateral compartments, and function was assessed by diuretic-inhibitable apicobasal volume transport assessed by weighing apical and basolateral supernatants. Cells were removed from the incubator and culture trays wrapped in sterile parafilm. Cell culture trays were placed in a conventional household 5^oC refrigerator for 24, 48, 72, and 96 hours (n=3 wells each condition) without any media exchanges. After the specified chill time, cell culture trays were removed from the refrigerator and chilled media exchanged for fresh warmed media. Cell culture plates were returned to culture at 37^oC and 5%CO₂/air. At 1, 12, and 40 days after chill exposure, cell barrier and transport function were again measured.

Results

Cells chilled for 24,48, and 72 hours maintained barrier integrity and transport function out to 40 days post chill. Cells chilled for 96 hours had an initial 28-fold increase in leak that persisted at lower levels to 40 days post exposure.

Conclusion

Renal tubule cells in bioartificial kidneys need to present a barrier to reabsorption of toxins and need to transport salt and water to concentrate filtered toxins. Primary renal tubule cells grown on permeable supports retained function and barrier even after 72 hours at 5C. Transport of bioartificial kidneys from manufacturing facilities to clinical sites may be possible with non-hazardous simple ice-water refrigeration.

Funding

• Private Foundation Support