Isolation of Primary Human Proximal Tubule Epithelial Cells and Their Use in Creating a Microphysiological Model of the Renal Proximal Tubule

Brooke Chalker; Yik Pui Tsang; Anish Mahadeo; Catherine K. Yeung; Jonathan Himmelfarb; Edward J. Kelly

doi:10.3791/67997

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Method Article

Isolation of Primary Human Proximal Tubule Epithelial Cells and Their Use in Creating a Microphysiological Model of the Renal Proximal Tubule

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DOI:

10.3791/67997

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May 9th, 2025

Brooke Chalker^*¹ , Yik Pui Tsang^*¹ , Anish Mahadeo¹ , Catherine K. Yeung²^,³ , Jonathan Himmelfarb⁴ , Edward J. Kelly¹^,³

¹Department of Pharmaceutics, School of Pharmacy, University of Washington, ²Department of Pharmacy, School of Pharmacy, University of Washington, ³Kidney Research Institute, Division of Nephrology, Department of Medicine, University of Washington, ⁴Barbara T. Murphy Division of Nephrology, Samuel Bronfman Department of Medicine, Icahn School of Medicine at Mount Sinai

^* These authors contributed equally

Summary

We present an optimized protocol for processing whole human kidneys to isolate and culture primary renal proximal tubule epithelial cells and the application of these cells in a three-dimensional, microfluidic, microphysiological platform to recapitulate the renal proximal tubule.

Abstract

Kidney disease affects over 850 million people globally, including 37 million Americans. Risk factors for chronic kidney disease include environmental influences, genetic predispositions, co-existing medical conditions, and a history of acute kidney injury. These factors often take months or years to develop, complicating longitudinal studies of disease etiology and pathophysiology. Advanced kidney models are needed to improve our understanding of disease mechanisms and enhance nephrotoxicity prediction in drug development. Proximal tubule epithelial cells (PTECs) in the kidney play a critical role in xenobiotic and toxin clearance as well as the reabsorption of essential nutrients. We have previously demonstrated that three-dimensional (3D) microphysiological system (MPS) platforms, populated with isolated primary PTECs, can be used to investigate renal drug interactions, assess nephrotoxicity of compounds, and predict drug clearance. Here, we present protocols for isolating and culturing primary PTECs from whole human kidneys and for seeding them into a 3D MPS platform that mimics in vivo renal physiology. This protocol enables long-term studies supporting PTEC viability, physiological morphology, and functional polarization of key transporter proteins in MPS devices for up to 6 months.

Introduction

The kidney plays a critical role in the clearance and elimination of a wide array of xenobiotics, toxins, and endogenous compounds from the body. This is achieved by filtering blood to remove waste products and by regulating electrolyte balance, fluid levels, and pH. Each human kidney contains around one million nephrons, the structural and functional units of the kidney¹. Within these nephrons, specialized epithelial cells in the proximal tubules, known as proximal tubule epithelial cells (PTECs), are responsible for reabsorbing essential molecules such as glucose, amino acids, and ions, as well as for secreting drug substrates and potentially....