Adaptive regulation of Na+-dependent phosphate transport in the bovine renal epithelial cell line NBL-1. Identification of the phosphate transporter as a 55-kDa glycoprotein

Hyperosmolarity induced an increase in Na(+)-dependent L-alanine uptake in confluent monolayers of the established renal epithelial cell line NBL-1. This induction was attributable to system A and was only seen when the cells had been previously deprived of amino acids in the culture medium to derepress system A activity. It was additive to the adaptive regulation induction, and both were inhibited by cycloheximide. However, the hyperosmolarity effect was inhibited by colcemid (an inhibitor of microtubular function), but adaptive regulation was not. Otherwise, when cell monolayers were incubated in a control medium, basal Na(+)-dependent L-alanine uptake mediated by system B0 decreased. The results of this study show that: (i) system A activity was not induced by cell shrinkage and subsequent swelling due to extracellular hyperosmolarity when cells were incubated in control medium; (ii) previous expression of system A activity induced by amino acid starvation seems to be a prerequisite for further induction due to hyperosmolarity; and (iii) the effects of adaptive regulation and hyperosmotic stress are mediated by different mechanisms.

Download Full-text

Electrophysiological Measurements of a Toad Renal Epithelial Cell Line (A6) as an Assay for Predicting Ocular Eye Irritancy

Alternatives to Laboratory Animals ◽

10.1177/026119299202000206 ◽

1992 ◽

Vol 20 (2) ◽

pp. 218-221

Author(s):

Henning F. Bjerregaard

Keyword(s):

Epithelial Cell ◽

Cell Line ◽

South African ◽

Mdck Cells ◽

Epithelial Cell Line ◽

Renal Epithelial Cell ◽

Electrophysiological Measurements ◽

Renal Epithelial Cell Line ◽

Darby Canine Kidney

An established epithelial cell line (A6) from a South African clawed toad (Xenopus laevis) kidney was used as a model for the corneal epithelium of the eye in order to determine ocular irritancy. When grown on Millipore filter inserts, A6 cells form a monolayer epithelium of high electrical resistance and generate a trans-epithelial potential difference. These two easily-measured electrophysiological endpoints showed a dose-related decrease after exposure for 24 hours to seven selected chemicals of different ocular irritancy potential. It was demonstrated that both trans-epithelial resistance and potential ranked closely with in vivo eye irritancy data and correlated well (r = 0.96) with loss of trans-epithelial impermeability of Madin-Darby canine kidney (MDCK) cells, detected by use of a fluorescein leakage assay.

Download Full-text

Polarized apical distribution of glycosyl-phosphatidylinositol-anchored proteins in a renal epithelial cell line.

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.85.24.9557 ◽

1988 ◽

Vol 85 (24) ◽

pp. 9557-9561 ◽

Cited By ~ 222

Author(s):

M. P. Lisanti ◽

M. Sargiacomo ◽

L. Graeve ◽

A. R. Saltiel ◽

E. Rodriguez-Boulan

Keyword(s):

Epithelial Cell ◽

Cell Line ◽

Epithelial Cell Line ◽

Renal Epithelial Cell ◽

Glycosyl Phosphatidylinositol ◽

Renal Epithelial Cell Line

Download Full-text

Expression of phosphoenolpyruvate carboxykinase (PEPCK) chimeras in renal epithelial cells. Retention of appropriate physiological responsiveness using enhancerless retroviral vectors

Biochemical Journal ◽

10.1042/bj2840725 ◽

1992 ◽

Vol 284 (3) ◽

pp. 725-732 ◽

Cited By ~ 2

Author(s):

A S Pollock ◽

D H Lovett

Keyword(s):

Epithelial Cell ◽

Cyclic Amp ◽

Cell Line ◽

Phosphoenolpyruvate Carboxykinase ◽

Large Fraction ◽

Expression System ◽

Retroviral Vectors ◽

Epithelial Cell Line ◽

Renal Epithelial Cell ◽

Renal Epithelial Cell Line

We used an enhancerless U3 mutant retroviral vector to deliver chimeras of the phosphoenolpyruvate carboxykinase (PEPCK) promoter region to a renal epithelial cell line capable of expressing PEPCK mRNA. Chimeras consisting of the PEPCK promoter and chloramphenicol acetyltransferase, neomycin phosphotransferase or human growth hormone genes were expressed after viral infection of the NRK52E renal epithelial cell line. Virus-delivered sequences in which the direction of PEPCK promoter transcription was antegrade to the normal direction of the long terminal repeat (LTR)-initiated transcription correctly upon stimulation with dexamethasone or 8-bromo cyclic AMP and upon lowering of the extracellular pH. Fluorescent primer extension in situ using primers specific for virus-delivered sequences of antegrade constructs indicated that a large fraction of NRK52E cells could be infected by co-cultivation with virus-producing psi-2 cells without G418 selection. Virus-delivered constructs whose orientation was opposite to that of the LTRs were expressed at very low levels, with transcripts detectable by PCR only in RNA from cyclic AMP-treated cells. Using reverse transcription/PCR, we demonstrated that the chimeric transcripts were from the internal PEPCK promoter rather than a functional or reconstituted Moloney LTR. PEPCK-reporter chimeras delivered by retroviral vectors demonstrated a level of expression more consistent with the level of expression of the native PEPCK gene than did transfected chimeras. This expression system should prove useful for studies of the physiological modulation of gene expression in renal tissues.

Download Full-text