scholarly journals A simple method for the isolation and detailed characterization of primary human proximal tubule cells for renal replacement therapy

2019 ◽  
Vol 43 (1) ◽  
pp. 45-57 ◽  
Author(s):  
Natalia Sánchez-Romero ◽  
Laura Martínez-Gimeno ◽  
Pedro Caetano-Pinto ◽  
Berta Saez ◽  
José Manuel Sánchez-Zalabardo ◽  
...  
Keyword(s):  
Proximal Tubule ◽  
Cancer Resistance ◽  
Simple Method ◽  
Waste Products ◽  
Proximal Tubule Cells ◽  
Tubule Cells ◽  
Glutamyl Transferase ◽  
Human Proximal Tubule

The main physiological functions of renal proximal tubule cells in vivo are reabsorption of essential nutrients from the glomerular filtrate and secretion of waste products and xenobiotics into urine. Currently, there are several established cell lines of human origin available as in vitro models of proximal tubule. However, these cells appeared to be limited in their biological relevance, because essential characteristics of the original tissue are lost once the cells are cultured. As a consequence of these limitations, primary human proximal tubule cells constitute a suitable and a biologically more relevant in vitro model to study this specific segment of the nephron and therefore, these cells can play an important role in renal regenerative medicine applications. Here, we describe a protocol to isolate proximal tubule cells from human nephrectomies. We explain the steps performed for an in-depth characterization of the cells, including the study of markers from others segments of the nephron, with the goal to determine the purity of the culture and the stability of proteins, enzymes, and transporters along time. The human proximal tubule cells isolated and used throughout this study showed many proximal tubule characteristics, including monolayer organization, cell polarization with the expression of tight junctions and primary cilia, expression of proximal tubule–specific proteins, such as megalin and sodium/glucose cotransporter 2, among others. The cells also expressed enzymatic activity for dipeptidyl peptidase IV, as well as for gamma glutamyl transferase 1, and expressed transporter activity for organic anion transporter 1, P-glycoprotein, multidrug resistance proteins, and breast cancer resistance protein. In conclusion, characterization of our cells confirmed presence of putative proximal tubule markers and the functional expression of multiple endogenous organic ion transporters mimicking renal reabsorption and excretion. These findings can constitute a valuable tool in the development of bioartificial kidney devices.

scholarly journals Transcriptomic characterization of short duration endoplasmic reticulum stress on cultured human proximal tubule cells

2014 ◽  
Vol 15 (Suppl 10) ◽  
pp. P5 ◽  
Author(s):  
Yan Zhang ◽  
Michelle Barati ◽  
Ignacio Munoz ◽  
Ming Li ◽  
Danny Wilkey ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Endoplasmic Reticulum Stress ◽  
Proximal Tubule ◽  
Short Duration ◽  
Proximal Tubule Cells ◽  
Tubule Cells ◽  
Human Proximal Tubule

Molecular Size and Origin Do Not Influence the Harmful Side Effects of Hydroxyethyl Starch on Human Proximal Tubule Cells (HK-2) In Vitro

2014 ◽  
Vol 119 (3) ◽  
pp. 570-577 ◽  
Author(s):  
Raphael R. Bruno ◽  
Winfried Neuhaus ◽  
Norbert Roewer ◽  
Christian Wunder ◽  
Martin A. Schick
Keyword(s):  
Side Effects ◽  
Proximal Tubule ◽  
Hydroxyethyl Starch ◽  
Molecular Size ◽  
Proximal Tubule Cells ◽  
Tubule Cells ◽  
Human Proximal Tubule

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

2007 ◽  
Vol 293 (2) ◽  
pp. F476-F485 ◽  
Author(s):  
Sara Terryn ◽  
François Jouret ◽  
Frank Vandenabeele ◽  
Inge Smolders ◽  
Marjan Moreels ◽  
...  
Keyword(s):  
Proximal Tubule ◽  
Short Circuit ◽  
Primary Cultures ◽  
Endocytic Pathway ◽  
Simple Method ◽  
Proximal Tubule Cells ◽  
Apical Side ◽  
Microscopic Evaluation ◽  
Tubule Cells ◽  
Glutamyl Transferase

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 γ-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.

scholarly journals SP781THE EFFECT OF IN VITRO TACROLIMUS EXPOSURE AND PHARMACOGENETIC VARIATION ON MULTILEVEL CYP3A5, ABCB1 EXPRESSION AND CTGF PRODUCTION IN HUMAN PROXIMAL TUBULE CELLS

2017 ◽  
Vol 32 (suppl_3) ◽  
pp. iii407-iii407
Author(s):  
Noel Knops ◽  
Yasaman Ramazani ◽  
Bert van den Heuvel ◽  
Elena Levtchenko ◽  
Dirk Kuypers
Keyword(s):  
Proximal Tubule ◽  
Proximal Tubule Cells ◽  
Tubule Cells ◽  
Human Proximal Tubule

scholarly journals Comparison of diglycolic acid exposure to human proximal tubule cells in vitro and rat kidneys in vivo

2017 ◽  
Vol 4 ◽  
pp. 342-347 ◽  
Author(s):  
Miriam E. Mossoba ◽  
Sanah Vohra ◽  
Howard Toomer ◽  
Shelia Pugh-Bishop ◽  
Zachary Keltner ◽  
...  
Keyword(s):  
Proximal Tubule ◽  
Acid Exposure ◽  
Proximal Tubule Cells ◽  
Diglycolic Acid ◽  
Tubule Cells ◽  
Human Proximal Tubule ◽  
Rat Kidneys

Freshly isolated primary human proximal tubule cells as an in vitro model for the detection of renal tubular toxicity

Toxicology ◽  
2020 ◽  
Vol 442 ◽  
pp. 152535 ◽  
Author(s):  
Piyush Bajaj ◽  
Git Chung ◽  
Keith Pye ◽  
Tomoya Yukawa ◽  
Akio Imanishi ◽  
...  
Keyword(s):  
Proximal Tubule ◽  
In Vitro Model ◽  
Proximal Tubule Cells ◽  
Tubular Toxicity ◽  
Tubule Cells ◽  
Vitro Model ◽  
Renal Tubular ◽  
Human Proximal Tubule

Nephrotoxicity Testing In Vitro: The Current Situation

1997 ◽  
Vol 25 (5) ◽  
pp. 497-503
Author(s):  
Jean-Paul Morin ◽  
Marc E. De Broe ◽  
Walter Pfaller ◽  
Gabriele Schmuck
Keyword(s):  
Proximal Tubule ◽  
Culture Media ◽  
Task Force ◽  
Primary Cultures ◽  
Phenotypic Expression ◽  
Transepithelial Transport ◽  
Specific Cell ◽  
Proximal Tubule Cells ◽  
Tubule Cells

An ECVAM task force on nephrotoxicity has been established to advise, in particular, on the follow-up to recommendations made in the ECVAM workshop report on nephrotoxicity testing in vitro. Since this workshop was held, in 1994, there have been several improvements in the techniques used. For example, the duration of renal slice viability, and the maintenance of functional activities in slices, have been improved by using dynamic incubation systems with higher oxygen tensions and more-appropriate cell culture media. Highly differentiated primary cultures of pig, human and rabbit proximal tubule cells have been established by using specific cell isolation procedures and/or selective culture media. To date, the most comparable phenotypic expression and transepithelial transport capacities to proximal tubules in vivo have been obtained with primary cultures of rabbit proximal tubule cells which are grown on bicompartmental supports; in this system, transepithelial substrate gradients are generated and the transepithelial transport of both organic anions and cations is highly active. This in vitro system has been selected by ECVAM for further evaluation and prevalidation. Industrial needs in the area of nephrotoxicity testing have been identified, and recommendations are made at the end of this report concerning possible future initiatives.

Sign in / Sign up

Export Citation Format

Share Document