A Novel Protocol for Culturing Polarized Proximal Tubular Epithelial Cells from Kidney Biopsies: Enhancing Platforms for Drug Excretion and Nephrotoxicity Studies

Abstract

The kidneys are integral to homeostasis but are susceptible to nephrotoxic compounds. Proximal tubular epithelial cells (PTECs) mediate drug metabolism and transport and are widely used in preclinical studies. However, commercial PTECs are limited in availability and physiological relevance. This study aimed to develop a novel, reliable protocol for isolating and culturing PTECs from human kidney biopsies. Primary PTECs were isolated from kidney biopsies of two patients (MFUM-RPTEC-1 and MFUM-RPTEC-2). Their morphology, population doubling time, transepithelial electrical resistance (TEER), and phenotypic markers were evaluated. Polarization and transporter expression were analyzed using cells cultured on Transwell inserts. Colonies formed within 24–48 h, with confluence reached by 8–10 days and dome (hemicyst) formation by day 13. TEER values peaked at 190 Ω/cm2 after 7–14 days, confirming tight junction formation. Immunostaining identified characteristic markers (e.g., SGLT2, OAT1/3, OCT2, P-gp, MRP4, MATE1, N-cadherin, ZO-1, CK-18). Cells cultured on Transwell plates exhibited native polarization, expressing transporters crucial for drug excretion on apical and basolateral surfaces. We present two robust protocols for isolating and characterizing PTECs, offering a scalable method to obtain functional, polarized cells from scarce biopsy material. The isolated PTECs, therefore, present a valuable platform for preclinical studies, especially for drug excretion testing through the expressed transporters. Drug competition for these transporters during tubular secretion is also a common cause of nephrotoxicity.

Keywords

• cell separation
• epithelial cells
• large-core needle biopsy
• phenotype
• renal excretion
• toxicity