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This Article Full Text Full Text (PDF) All Versions of this Article: 293/2/F476    most recent 00363.2006v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in ISI Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Terryn, S. Articles by Kerkhove, E. V. Search for Related Content PubMed PubMed Citation Articles by Terryn, S. Articles by Kerkhove, E. V.

A primary culture of mouse proximal tubular cells, established on collagen-coated membranes

Sara Terryn,^1,4 Franois Jouret,⁵ Frank Vandenabeele,^2,3 Inge Smolders,^1,3 Marjan Moreels,^2,3 Olivier Devuyst,³ Paul Steels,^1,3 and Emmy Van Kerkhove^1,4

¹Laboratory of Cell Physiology and ²Laboratory of Histology, Hasselt University and Transnationale Universiteit Limburg, ³Biomedical Institute and ⁴Centre for Environmental Sciences, Diepenbeek, Belgium; and ³Division of Nephrology, Université Catholique de Louvain, Brussels, Belgium

Submitted 11 September 2006 ; accepted in final form 28 April 2007

A simple method is described to establish primary cultures of kidney proximal tubule cells (PTC) on membranes. The permeable membranes represent a unique culture surface, allowing a high degree of differentiation since both apical and basolateral membranes are accessible for medium. Proximal tubule (PT) segments from collagenase-digested mouse renal cortices were grown for 7 days, by which time cells were organized as a confluent monolayer. Electron microscopic evaluation revealed structurally polarized epithelial cells with numerous microvilli, basolateral invaginations, and apical tight junctions. Immunoblotting for markers of distinct parts of the nephron demonstrated that these primary cultures only expressed PT-specific proteins. Moreover immunodetection of distinct components of the receptor-mediated endocytic pathway and uptake of FITC-albumin indicated that these cells expressed a functional endocytotic apparatus. In addition, primary cultures possessed the PT brush-border enzymes, alkaline phosphatase, and -glutamyl-transferase, and a phloridzin-sensitive sodium-dependent glucose transport at their apical side. Electrophysiological measurements show that the primary cultured cells have a low transepithelial resistance and high short-circuit current that was completely carried by Na⁺ similar to a leaky epithelium like proximal tubule cells. This novel method established well-differentiated PTC cultures.

aquaporin; electrophysiological characteristics; phloridzin; receptor-mediated endocytosis; sodium-dependent glucose transport; Ussing chamber