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Drug transporter expression profiling in a three-dimensional kidney proximal tubule in vitro nephrotoxicity model

  • Ion channels, receptors and transporters
  • Published:
Pflügers Archiv - European Journal of Physiology Aims and scope Submit manuscript

Abstract

Given currently poor toxicity translational predictions for drug candidates, improved mechanistic understanding underlying nephrotoxicity and drug renal clearance is needed to improve drug development and safety screening. Therefore, better relevant and well-characterized in vitro screening models are required to reliably predict human nephrotoxicity. Because kidney proximal tubules are central to active drug uptake and secretion processes and therefore to nephrotoxicity, this study acquired regio-specific expression data from recently reported primary proximal tubule three-dimensional (3D) hyaluronic acid gel culture and non-gel embedded cultured murine proximal tubule suspensions used in nephrotoxicity assays. Quantitative assessment of the mRNA expression of 21 known kidney tubule markers and important proximal tubule transporters with known roles in drug transport was obtained. Asserting superior gene expression levels over current commonly used two-dimensional (2D) kidney cell culture lines was the study objective. Hence, we compare previously published gel-based 3D proximal tubule fragment culture and their non-gel suspensions for up to 1 week. We demonstrate that 3D tubule culture exhibits superior gene expression levels and profiles compared to published commonly used 2D kidney cell lines (Caki-1 and HK-2) in plastic plate monocultures. Additionally, nearly all tested genes retain mRNA expression after 7 days in both proximal tubule cultures, a limitation of 2D cell culture lines. Importantly, gel presence is shown not to interfere with the gene expression assay. Western blots confirm protein expression of OAT1 and 3 and OCT2. Functional transport assays confirm their respective transporter functions in vitro. Overall, results validate retention of essential toxicity-relevant transporters in this published 3D proximal tubule model over conventional 2D kidney cell cultures, producing opportunities for more reliable, sensitive, and comprehensive drug toxicity studies relevant to drug development and nephrotoxicity goals.

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Acknowledgments

This work was supported by the AstraZeneca UK Limited, London, England. We thank Dr. M. Wagoner at AstraZeneca Pharmaceuticals, USA, for valuable discussions. DWG is grateful for support from the George S. and Dolores Doré Eccles Foundation (USA).

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  1. Department of Pharmaceutics and Pharmaceutical Chemistry, University of Utah, Salt Lake City, UT, 84112-5820, USA

    Dorina Diekjürgen & David W. Grainger

  2. Department of Bioengineering, University of Utah, Salt Lake City, UT, 84112-5820, USA

    David W. Grainger

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  1. Dorina Diekjürgen
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  2. David W. Grainger
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Correspondence to David W. Grainger.

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Diekjürgen, D., Grainger, D.W. Drug transporter expression profiling in a three-dimensional kidney proximal tubule in vitro nephrotoxicity model. Pflugers Arch - Eur J Physiol 470, 1311–1323 (2018). https://doi.org/10.1007/s00424-018-2150-z

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  • DOI: https://doi.org/10.1007/s00424-018-2150-z

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