Vasopressin stimulates DNA synthesis and ion transport in quiescent epithelial cells

1985 ◽  
Vol 249 (3) ◽  
pp. C267-C270 ◽  
Author(s):  
V. M. Reznik ◽  
R. J. Shapiro ◽  
S. A. Mendoza
Keyword(s):  
Epithelial Cells ◽  
Dna Synthesis ◽  
Ion Transport ◽  
Growth Regulation ◽  
Mdck Cells ◽  
Subsequent Increase ◽  
Low Concentrations ◽  
Pump Activity ◽  
Monovalent Ion ◽  
Na Uptake

The mitogenic effect of vasopressin was studied in subconfluent quiescent renal epithelial cells (MDCK). Vasopressin stimulated DNA synthesis in the presence of low concentrations of serum. Vasopressin increased the entry of Na into the cells and increased ouabain-sensitive 86Rb uptake, a measure of Na-K pump activity. Because the activity of the Na-K pump in MDCK cells is steeply dependent on intracellular Na, it is likely that stimulation of the Na-K pump by vasopressin was mediated by the increase in Na entry into the cells. Thus both serum and vasopressin stimulate Na uptake and Na-K pump activity in quiescent MDCK cells with a subsequent increase in DNA synthesis. It is concluded that growth regulation in epithelial cells may be mediated in part by changes in monovalent ion transport.

Low efficiency of functional translation of ouabain-resistant α2-Na+,K+-ATPase mRNA in C7-MDCK epithelial cells

2012 ◽  
Vol 90 (1) ◽  
pp. 83-88
Author(s):  
Olga A. Akimova ◽  
James Van Huysse ◽  
Johanne Tremblay ◽  
Sergei N. Orlov
Keyword(s):  
Epithelial Cells ◽  
Mdck Cells ◽  
Protein A ◽  
Cytotoxic Action ◽  
Atpase Subunit ◽  
Transfected Cells ◽  
Subunit Mrna ◽  
Pump Activity ◽  
Low Efficiency

Na+,K+-ATPase is a heterodimer consisting of catalytic α1–α4 and regulatory β1–β3 subunits. Recently, we reported that transfection with ouabain-resistant α1R-Na+,K+-ATPase rescues renal epithelial C7-MDCK cells exclusively expressing the ouabain-sensitive α1S-isoform from the cytotoxic action of ouabain. To explore the role of α2 subunit in ion transport and cytotoxic action of ouabain, we compared the effect of ouabain on K+ (86Rb) influx and the survival of ouabain-treated C7-MDCK cells stably transfected with α1R- and α2R-Na+,K+-ATPase. α2R mRNA in transfected cells was ∼8-fold more abundant than α1R mRNA, whereas immunoreactive α2R protein content was 5-fold lower than endogenous α1S protein. A concentration of 10 µmol/L ouabain led to complete inhibition of 86Rb influx both in mock- and α2R-transfected cells, whereas maximal inhibition of 86Rb influx in α1R-transfectd cells was observed at 1000 µmol/L ouabain. In contrast to the massive death of mock- and α2R-transfected cells exposed to 3 µmol/L ouabain , α1R-cells survived after 24 h incubation with 1000 µmol/L ouabain. Thus, our results show that unlike α1R, the presence of α2R-Na+,K+-ATPase subunit mRNA and immunoreactive protein does not contribute to Na+/K+ pump activity, and does not rescue C7-MDCK cells from the cytotoxic action of ouabain. Our results also suggest that the lack of impact of transfected α2-Na+,K+-ATPase on Na+/K+ pump activity and cell survival can be attributed to the low efficiency of its translation and (or) delivery to the plasma membrane of renal epithelial cells.

Urea selectively induces DNA synthesis in renal epithelial cells

1993 ◽  
Vol 264 (4) ◽  
pp. F601-F607 ◽  
Author(s):  
D. M. Cohen ◽  
S. R. Gullans
Keyword(s):  
Epithelial Cells ◽  
Dna Synthesis ◽  
Thymidine Incorporation ◽  
Mdck Cells ◽  
Mrna Level ◽  
Cell Number ◽  
Total Protein Content ◽  
Cell Cycle Distribution ◽  
Promoting Effect ◽  
Renal Epithelial Cells

Hyperosmotic stress with the functionally impermeant solute NaCl has been shown by us and others to inhibit cell growth and DNA synthesis. Several lines of evidence suggest that urea, the other principal renal medullary solute, may exert a growth-promoting effect on renal epithelial cells. Among these is the finding that urea upregulates expression at the mRNA level of two growth-associated immediate-early genes, Egr-1 and c-fos. In the present study, urea, in concentrations characteristic of the renal medulla, increased [3H]thymidine incorporation approximately threefold in confluent, growth-suppressed Madin-Darby canine kidney (MDCK) cells, whereas another readily membrane-permeant solute, glycerol, did not. Urea also overcame the inhibitory effect of hyperosmotic NaCl on DNA synthesis. The urea-induced increase in [3H]thymidine incorporation was also evident in the renal epithelial LLC-PK1 cell line, but not in renal nonepithelial and epithelial nonrenal cell types examined. In addition, it was associated with a 15% increase in total DNA content measured fluorometrically at 24 h of treatment. There was, however, no associated increase in cell proliferation as measured by cell number, total protein content, or cell cycle distribution. Urea also failed to induce polyploidy or aneuploidy. Therefore cells of renal epithelial origin may be uniquely capable of responding to hyperosmotic urea with increased DNA synthesis through an undefined and potentially novel mechanism.

scholarly journals Early events elicited by bombesin and structurally related peptides in quiescent Swiss 3T3 cells. II. Changes in Na+ and Ca2+ fluxes, Na+/K+ pump activity, and intracellular pH.

1986 ◽  
Vol 102 (6) ◽  
pp. 2223-2233 ◽  
Author(s):  
S A Mendoza ◽  
J A Schneider ◽  
A Lopez-Rivas ◽  
J W Sinnett-Smith ◽  
E Rozengurt
Keyword(s):  
Ion Transport ◽  
Intracellular Ph ◽  
Free Calcium ◽  
3T3 Cells ◽  
Phosphotransferase System ◽  
Phorbol Dibutyrate ◽  
Protein Kinase C Activity ◽  
Pump Activity ◽  
Monovalent Ion ◽  
Swiss 3T3

The amphibian tetradecapeptide, bombesin, and structurally related peptides caused a marked increase in ouabain-sensitive 86Rb+ uptake (a measure of Na+/K+ pump activity) in quiescent Swiss 3T3 cells. This effect occurred within seconds after the addition of the peptide and appeared to be mediated by an increase in Na+ entry into the cells. The effect of bombesin on Na+ entry and Na+/K+ pump activity was concentration dependent with half-maximal stimulation occurring at 0.3-0.4 nM. The structurally related peptides litorin, gastrin-releasing peptide, and neuromedin B also stimulated ouabain-sensitive 86Rb+ uptake; the relative potencies of these peptides in stimulating the Na+/K+ pump were comparable to their potencies in increasing DNA synthesis (Zachary, I., and E. Rozengurt, 1985, Proc. Natl. Acad. Sci. USA., 82:7616-7620). Bombesin increased Na+ influx, at least in part, through an Na+/H+ antiport. The peptide augmented intracellular pH and this effect was abolished in the absence of extracellular Na+. In addition to monovalent ion transport, bombesin and the structurally related peptides rapidly increased the efflux of 45Ca2+ from quiescent Swiss 3T3 cells. This Ca2+ came from an intracellular pool and the efflux was associated with a 50% decrease in total intracellular Ca2+. The peptides also caused a rapid increase in cytosolic free calcium concentration. Prolonged pretreatment of Swiss 3T3 cells with phorbol dibutyrate, which causes a loss of protein kinase C activity (Rodriguez-Pena, A., and E. Rozengurt, 1984, Biochem. Biophys. Res. Commun., 120:1053-1059), greatly decreased the stimulation of 86Rb+ uptake and Na+ entry by bombesin implicating this phosphotransferase system in the mediation of part of these responses to bombesin. Since some activation of monovalent ion transport by bombesin was seen in phorbol dibutyrate-pretreated cells, it is likely that the peptide also stimulates monovalent ion transport by a second mechanism.

Possible role of ROS as mediators of hypoxia-induced ion transport inhibition of alveolar epithelial cells

2000 ◽  
Vol 278 (4) ◽  
pp. L640-L648 ◽  
Author(s):  
Wolf Heberlein ◽  
Ralf Wodopia ◽  
Peter Bärtsch ◽  
Heimo Mairbäurl
Keyword(s):  
Epithelial Cells ◽  
Ion Transport ◽  
A549 Cells ◽  
Alveolar Epithelial Cells ◽  
Transport Activity ◽  
Excitable Cells ◽  
Alveolar Epithelial ◽  
Oxygen Sensitive ◽  
Na Uptake

In oxygen-sensitive excitable cells, responses to hypoxia are initiated by membrane depolarization due to closing of the K channels that is thought to be mediated by a decrease in reactive oxygen species (ROS). Because the mechanisms of hypoxic inhibition of ion transport of alveolar epithelial cells (Planes C, Friedlander G, Loiseau A, Amiel C, and Clerici C. Am J Physiol Lung Cell Mol Physiol 271: L70–L78, 1996; Mairbäurl H, Wodopia R, Eckes S, Schulz S, and Bärtsch P. Am J Physiol Lung Cell Mol Physiol 273: L797–L806, 1997) are not yet understood, we tested the possible involvement of a hypoxia-induced change in ROS that might control transport activity. Transport was measured as86Rb and22Na uptake in A549 cells exposed to normoxia, hyperoxia, or hypoxia together with ROS donors and scavengers. H2O2< 1 mM did not affect transport, whereas 1 mM H2O2activated22Na uptake (+200%) but inhibited86Rb uptake (−30%). Also hyperoxia, aminotriazole plus menadione, and diethyldithiocarbamate inhibited86Rb uptake. N-acetyl-l-cysteine, diphenyleneiodonium, and tetramethylpiperidine- N-oxyl, used to reduce ROS, inhibited86Rb uptake, thus mimicking the hypoxic effects, whereas deferoxamine, superoxide dismutase, and catalase were ineffective. Also, hypoxic effects on ion transport were not prevented in the presence of H2O2, diethyldithiocarbamate, and N-acetyl-l-cysteine. These results indicate that ion transport of A549 cells is significantly affected by decreasing or increasing cellular ROS levels and that it is possible that certain species of ROS might mediate the hypoxic effects on ion transport of alveolar epithelial cells.

scholarly journals New Trimethoprim-Like Molecules: Bacteriological Evaluation and Insights into Their Action

Antibiotics ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 709
Author(s):  
Marta Jorba ◽  
Marina Pedrola ◽  
Ouldouz Ghashghaei ◽  
Rocío Herráez ◽  
Lluis Campos-Vicens ◽  
...  
Keyword(s):  
Inhibitory Concentration ◽  
Efflux Pump ◽  
Synergistic Effects ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Detailed Characterization ◽  
Low Concentrations ◽  
Pump Activity ◽  
Bacteriological Evaluation ◽  
Kinetics Of

This work reports a detailed characterization of the antimicrobial profile of two trimethoprim-like molecules (compounds 1a and 1b) identified in previous studies. Both molecules displayed remarkable antimicrobial activity, particularly when combined with sulfamethoxazole. In disk diffusion assays on Petri dishes, compounds 1a and 1b showed synergistic effects with colistin. Specifically, in combinations with low concentrations of colistin, very large increases in the activities of compounds 1a and 1b were determined, as demonstrated by alterations in the kinetics of bacterial growth despite only slight changes in the fractional inhibitory concentration index. The effect of colistin may be to increase the rate of antibiotic entry while reducing efflux pump activity. Compounds 1a and 1b were susceptible to extrusion by efflux pumps, whereas the inhibitor phenylalanine arginyl β-naphthylamide (PAβN) exerted effects similar to those of colistin. The interactions between the target enzyme (dihydrofolate reductase), the coenzyme nicotinamide adenine dinucleotide phosphate (NADPH), and the studied molecules were explored using enzymology tools and computational chemistry. A model based on docking results is reported.

Aberrant responses to growth-regulatory signals by variant kidney epithelial cells

1988 ◽  
Vol 254 (5) ◽  
pp. F747-F753
Author(s):  
M. M. Walsh-Reitz ◽  
R. I. Feldman ◽  
F. G. Toback
Keyword(s):  
Epithelial Cells ◽  
Growth Regulation ◽  
Growth Inhibitor ◽  
Soft Agar ◽  
Isoenzyme Analysis ◽  
Variant Cell ◽  
Kidney Epithelial Cells ◽  
Potent Growth ◽  
Potent Growth Inhibitor ◽  
Low K

Cultures that achieved a higher cell density than expected were noted during study of growth regulation in monkey kidney epithelial cells of the BSC-1 line. Multiplication of the variant cells was accelerated, compared with parental cells, as the cultures approached confluence. Cytogenetic analysis, immunofluorescence antibody reactions with specific monkey serum, isoenzyme analysis, microbiological studies, and lack of growth in soft agar indicated that the variant cells were not a contaminating cell type, lacked new isoenzymes, were free of microbial contamination, and were not transformed. Confluent variant cultures did not respond to a purified growth inhibitor protein produced by BSC-1 cells that inhibits multiplication and reduces cell Na content in subconfluent variant and parental cells. Vasopressin, which is a mitogen for parental cells, was a potent growth inhibitor for confluent cultures of variant cells. Low-K or high-Na media, which stimulate proliferation of parental cells, had no effect on growth of the variant cell line. These results suggest that enhanced multiplication of the variant cells is mediated by altered signal transduction pathways and/or receptors for growth-regulatory molecules.

Model translocators for divalent and monovalent ion transport in phospholipid membranes

1974 ◽  
Vol 18 (1) ◽  
pp. 187-199 ◽  
Author(s):  
H. Célis ◽  
S. Estrada-O ◽  
M. Montal
Keyword(s):  
Ion Transport ◽  
Phospholipid Membranes ◽  
Monovalent Ion

scholarly journals Atypical Protein Kinase C Is Involved in the Evolutionarily Conserved Par Protein Complex and Plays a Critical Role in Establishing Epithelia-Specific Junctional Structures

2001 ◽  
Vol 152 (6) ◽  
pp. 1183-1196 ◽  
Author(s):  
Atsushi Suzuki ◽  
Tomoyuki Yamanaka ◽  
Tomonori Hirose ◽  
Naoyuki Manabe ◽  
Keiko Mizuno ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cell Polarity ◽  
Protein Complex ◽  
Mdck Cells ◽  
Critical Role ◽  
Dominant Negative ◽  
Dominant Negative Mutant ◽  
Surface Polarity ◽  
C Elegans ◽  
Evolutionarily Conserved

We have previously shown that during early Caenorhabditis elegans embryogenesis PKC-3, a C. elegans atypical PKC (aPKC), plays critical roles in the establishment of cell polarity required for subsequent asymmetric cleavage by interacting with PAR-3 [Tabuse, Y., Y. Izumi, F. Piano, K.J. Kemphues, J. Miwa, and S. Ohno. 1998. Development (Camb.). 125:3607–3614]. Together with the fact that aPKC and a mammalian PAR-3 homologue, aPKC-specific interacting protein (ASIP), colocalize at the tight junctions of polarized epithelial cells (Izumi, Y., H. Hirose, Y. Tamai, S.-I. Hirai, Y. Nagashima, T. Fujimoto, Y. Tabuse, K.J. Kemphues, and S. Ohno. 1998. J. Cell Biol. 143:95–106), this suggests a ubiquitous role for aPKC in establishing cell polarity in multicellular organisms. Here, we show that the overexpression of a dominant-negative mutant of aPKC (aPKCkn) in MDCK II cells causes mislocalization of ASIP/PAR-3. Immunocytochemical analyses, as well as measurements of paracellular diffusion of ions or nonionic solutes, demonstrate that the biogenesis of the tight junction structure itself is severely affected in aPKCkn-expressing cells. Furthermore, these cells show increased interdomain diffusion of fluorescent lipid and disruption of the polarized distribution of Na+,K+-ATPase, suggesting that epithelial cell surface polarity is severely impaired in these cells. On the other hand, we also found that aPKC associates not only with ASIP/PAR-3, but also with a mammalian homologue of C. elegans PAR-6 (mPAR-6), and thereby mediates the formation of an aPKC-ASIP/PAR-3–PAR-6 ternary complex that localizes to the apical junctional region of MDCK cells. These results indicate that aPKC is involved in the evolutionarily conserved PAR protein complex, and plays critical roles in the development of the junctional structures and apico-basal polarization of mammalian epithelial cells.

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