Substrate Elasticity Governs Differentiation of Renal Tubule Cells in Prolonged Culture

Abstract

End-stage renal disease afflicts ∼750,000 Americans and claims >100,000 lives annually in the United States. Kidney transplantation is associated with longest survival and least cost but is limited by scarcity of donor organs. The balance of patients are treated with dialysis, a cumbersome, morbid, and expensive procedure. Each hemodialysis treatment consumes in excess of 160 liters of water and anchors the patient to a machine for 12–15 h per week. Cultured tubule cells can reduce the obligate fluid requirements of a bioengineered artificial kidney by concentrating wastes and reabsorbing filtered salt and water. Primary tubule epithelial cells rapidly dedifferentiate in culture and form a flattened epithelium lacking the brush border essential to apicobasal transport. We hypothesized that substrate mechanical properties have a strong influence on differentiation in primary cell culture. We cultured primary renal tubule cells on polyacrylamide hydrogels of varying elasticity and measured expression of key transporter proteins essential to renal tubule cell function. Primary tubule cells cultured on soft substrates for extended periods showed increased expression of key transporters characteristic of differentiated proximal tubule cells. These data support the hypothesis that scaffold elasticity is a critical factor in cell culture, and, unexpectedly, that prolonged culture of primary cells was essential to observing this difference.

Impact Statement

Successful clinical tissue engineering requires functional fidelity of the cultured cell to its in vivo counterpart, but this has been elusive in renal tissue engineering. Typically, renal proximal tubule cells in culture have a flattened morphology and do not express key transporters essential to their function. In this article, we show for the first time that in vitro substrate mechanical properties dictate differentiation of cultured renal proximal tubule cells. Remarkably, this effect was only discernable after 4 weeks in culture, longer than usually reported for this cell type. These results demonstrate a new tunable parameter to optimize cell differentiation in renal tissue engineering.