Increased functional differentiation of rabbit proximal tubule cells cultured in glucose-free media

1992 ◽  
Vol 263 (1) ◽  
pp. F152-F162
Author(s):  
A. Blais ◽  
F. Jalal ◽  
P. Crine ◽  
J. Paiement ◽  
A. Berteloot
Keyword(s):  
Proximal Tubule ◽  
Brush Border ◽  
Brush Border Membrane ◽  
Primary Cultures ◽  
Morphological Differentiation ◽  
Functional Expression ◽  
Functional Differentiation ◽  
Rabbit Kidney ◽  
Proximal Tubule Cells ◽  
Tubule Cells

We have determined the influence of glucose (Glc)-free medium on the growth and differentiation of rabbit kidney proximal tubule cells (PTC) in primary cultures. The specific growth rates and the protein-to-volume ratios are shown to be independent of the culture conditions. In contrast, the functional expression of four brush-border membrane enzyme markers was found to decline steadily and in the same way from day 3 in culture up to late confluence in Glc-containing medium, and different evolution patterns and high expression levels were observed up to confluence in a Glc-free glutamine (Gln)-supplemented medium. Electron microscopy clearly showed, however, that the functional and morphological differentiation of the brush-border membrane is not correlated. Finally, by use of an indirect immunofluorescent technique in combination with flow cytometry, it is demonstrated that confluent cells grown in Glc and Gln media form homogeneous cell populations of PTC. It is thus concluded that the functional differentiation of rabbit kidney PTC in primary cultures is strongly dependent upon the energy source present in the culture medium.

Immunolocalization of NHE8 in rat kidney

2005 ◽  
Vol 288 (3) ◽  
pp. F530-F538 ◽  
Author(s):  
Sunita Goyal ◽  
SueAnn Mentone ◽  
Peter S. Aronson
Keyword(s):  
Proximal Tubule ◽  
Brush Border ◽  
Brush Border Membrane ◽  
Rat Kidney ◽  
Surface Membrane ◽  
Proximal Tubules ◽  
Intense Staining ◽  
Proximal Tubule Cells ◽  
Nephron Segment ◽  
Tubule Cells

In situ hybridization studies demonstrated that Na+/H+ exchanger NHE8 is expressed in kidney proximal tubules. Although membrane fractionation studies suggested apical brush-border localization, precise membrane localization could not be definitively established. The goal of the present study was to develop isoform-specific NHE8 antibodies as a tool to directly establish the localization of NHE8 protein in the kidney by immunocytochemistry. Toward this goal, two sets of antibodies that label different NHE8 epitopes were developed. Monoclonal antibody 7A11 and polyclonal antibody Rab65 both specifically labeled NHE8 by Western blotting as well as by immunofluorescence microscopy. The immunolocalization pattern in the kidney seen with both antibodies was the same, thereby validating NHE8 specificity. In particular, NHE8 expression was observed on the apical brush-border membrane of all proximal tubules from S1 to S3. The most intense staining was evident in proximal tubules in the deeper cortex and medulla with a significant but somewhat weaker staining in superficial proximal tubules. Colocalization studies with γ-glutamyltranspeptidase and megalin indicated expression of NHE8 on both the microvillar surface membrane and the coated-pit region of proximal tubule cells, suggesting that NHE8 may be subject to endocytic retrieval and recycling. Although colocalizing in the proximal tubule with NHE3, no significant alteration in NHE8 protein expression was evident in NHE3-null mice. We conclude that NHE8 is expressed on the apical brush-border membrane of proximal tubule cells, where it may play a role in mediating or regulating ion transport in this nephron segment.

scholarly journals Study of the mechanism by which the Na+-Pi co-transporter of mouse kidney proximal-tubule cells adjusts to phosphate depletion

1988 ◽  
Vol 252 (1) ◽  
pp. 105-109 ◽  
Author(s):  
M Jahan ◽  
P J Butterworth
Keyword(s):  
Proximal Tubule ◽  
Brush Border ◽  
Brush Border Membrane ◽  
Mouse Kidney ◽  
Transport Activity ◽  
Normal Medium ◽  
Proximal Tubule Cells ◽  
Tubule Cells ◽  
Kinetic Investigations ◽  
Stimulation Of

1. Proximal-tubule cells isolated from mouse kidney after digestion with collagenase take up Pi by an Na+-dependent and saturable process mediated by the Na+-Pi co-transporter of the brush-border membrane. 2. Pi depletion of the cells is accompanied by a stimulation of Pi-transport activity. Kinetic investigations reveal that Vmax. is increased by 90% and Km decreased by 50% after Pi depletion. Transport activity returns to normal values after incubation for 30 min at 37 degrees C of Pi-depleted cells in normal medium containing 1 mM-Pi, but the fall in transport activity under these conditions is inhibited by colchicine. 3. The energy of activation of Na+-Pi co-transport activity of depleted cells differs greatly from that found for normal replete cells. 4. The results provide evidence that stimulation of transport by Pi depletion arises from an increase in the number of carrier sites in the brush-border membrane. Additionally, changes in the properties of the transporter occur which may reflect altered phospholipid-carrier-protein interaction in the Pi-depleted condition.

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.

Primary cultures of renal proximal tubule cells derived from individuals with primary hyperoxaluria

2009 ◽  
Vol 37 (3) ◽  
pp. 127-132 ◽  
Author(s):  
Karen L. Price ◽  
Sally-Anne Hulton ◽  
William G. van’t Hoff ◽  
John R. Masters ◽  
Gill Rumsby
Keyword(s):  
Proximal Tubule ◽  
Primary Cultures ◽  
Primary Hyperoxaluria ◽  
Renal Proximal Tubule ◽  
Proximal Tubule Cells ◽  
Tubule Cells ◽  
Renal Proximal Tubule Cells
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