Heat shock-induced protection and enhancement of Na+-glucose cotransport by LLC-PK1 monolayers

1997 ◽  
Vol 273 (4) ◽  
pp. F530-F537 ◽  
Author(s):  
Caroline R. Sussman ◽  
J. Larry Renfro
Keyword(s):  
Heat Shock ◽  
Cell Survival ◽  
Surface Area ◽  
Membrane Surface ◽  
Epithelial Cell Line ◽  
Transport Capacity ◽  
Renal Epithelial Cell ◽  
Membrane Surface Area ◽  
Luminal Membrane ◽  
Renal Epithelial Cell Line

Monolayers of the porcine-derived renal epithelial cell line, LLC-PK1, were used to characterize the effects of heat stress on Na+-glucose cotransport. Transepithelial current dependent on 5 mM glucose ( I Glc), phloridzin-sensitive current ( I phz), and total transepithelial current ( I total) were measured as indicators of Na+-glucose cotransport. Severe heat shock (SHS; 45°C for 1 h, then 37°C for measurements) decreased transepithelial electrical resistance (TER), I Glc, I phz, and I total50–70%. Mild heat shock (MHS; 42°C for 3 h, then 37°C for 12 h) induced accumulation of 72-kDa heat shock protein (HSP-72), decreased damage to TER from SHS, and prevented damage to I Glc, I phz, and I total. Kinetic analysis showed that SHS damaged and MHS protected total Na+-glucose transport capacity ( V max of I Glc). MHS alone increased TER (50%), I Glc (20%), I total (20%), and V max of I Glc (25%). On enhancement of the Na+ gradient by depletion of intracellular Na+, MHS increased I Glc 50% and had no effect on transepithelial Na+-dependent sulfate reabsorptive flux measured concurrently or in Na+-replete tissues. These effects of MHS were not reflected in effects on cell survival or luminal membrane surface area as indicated by lactate dehydrogenase or alkaline phosphatase release. In conclusion, HSP-72-inducing heat treatment both protected and enhanced Na+-glucose cotransport independently of the luminal membrane Na+ gradient and selectively with respect to effects on TER, reabsorptive sulfate transport, cell survival, and luminal membrane surface area.

Gentamicin uptake by LLCPK1 cells: effect of intracellular and extracellular pH changes

1998 ◽  
Vol 76 (2) ◽  
pp. 155-160 ◽  
Author(s):  
Eric Chi-Chung Lui ◽  
Reina Bendayan
Keyword(s):  
Organic Cation ◽  
Extracellular Ph ◽  
Epithelial Cell Line ◽  
Renal Epithelial Cell ◽  
Ph Changes ◽  
Luminal Membrane ◽  
Different Temperatures ◽  
Renal Proximal Tubular Cells ◽  
Renal Epithelial Cell Line ◽  
Renal Proximal Tubular

The mechanisms by which aminoglycosides are transported across the luminal membrane of renal proximal tubular cells remain unclear. A luminal organic cation/H+ exchange as well as an adsorptive endocytosis membrane process has been proposed to be involved in gentamicin renal accumulation. The objectives of this work were to explore further the effects of intracellular and extracellular pH changes on gentamicin uptake. [3H]Gentamicin uptake by a continuous renal epithelial cell line, LLCPK1, grown as a monolayer on an impermeable surface was measured at different temperatures and pH conditions and in the presence of various inhibitors. Uptake of gentamicin was found to be carrier mediated (Km = 1.26 ± 0.22 mM, Vmax = 289 ± 27 pmol ·mg-1 ·min-1), energy dependent (inhibited in part by sodium azide), and temperature dependent (37°C > 4°C). Fifteen-minute gentamicin (10 µM) uptake was inhibited by 1 mM of the organic cations cimetidine (61.0%), quinidine (73.5%), quinine (68.6%), and verapamil (61.5%). More importantly, while an outwardly directed proton gradient did not have a significant effect on gentamicin uptake, extracellular acidification (pH 6.5), which leads to a higher degree of gentamicin ionization, significantly enhanced gentamicin uptake by LLCPK1 monolayer cells. These results suggest that the luminal organic cation/H+ exchanger is not involved in gentamicin uptake by renal cultured epithelial cells. Rather, the cationic charge of gentamicin appears to be one of the primary determinants for renal luminal uptake.Key words: gentamicin, LLCPK1 cells, pH, nephrotoxicity.

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

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.

scholarly journals Urotensin II is an Autocrine/Paracrine Growth Factor for the Porcine Renal Epithelial Cell Line, LLCPK1

Endocrinology ◽  
2003 ◽  
Vol 144 (5) ◽  
pp. 1825-1831 ◽  
Author(s):  
Mika Matsushita ◽  
Masayoshi Shichiri ◽  
Nozomi Fukai ◽  
Naoko Ozawa ◽  
Takanobu Yoshimoto ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Growth Factor ◽  
Kinase Inhibitor ◽  
Epithelial Cell Line ◽  
Urotensin Ii ◽  
Renal Epithelial Cell ◽  
Renal Epithelial Cells ◽  
Kinase C ◽  
Renal Epithelial Cell Line ◽  
Paracrine Growth Factor

Urotensin-II (UII), a cyclic dodecapeptide with potent cardiovascular effects, has recently been shown to be abundantly expressed in the human kidney and excreted in human urine. To investigate whether UII acts as an autocrine/paracrine growth factor for renal epithelial cells, we have studied the effects of human UII (hUII) on DNA synthesis, cytosolic free Ca2+ concentration ([Ca2+]i), ERK activation, and protooncogene (c-myc) expression in a porcine renal epithelial cell line (LLCPK1). hUII stimulated [3H]thymidine uptake into quiescent cells in a dose-dependent manner (10−9 to 10−7m); this effect was inhibited by a protein kinase C inhibitor (GF109203X), a MAPK kinase inhibitor (PD98059), and a calcium channel blocker (nicardipine). Neither phosphatidyl inositol-3 kinase inhibitors (LY294002, wortmannin) nor p38 kinase inhibitor (SB203580) affected the hUII-induced DNA syntheses. hUII rapidly (within 5 min) and dose-dependently (10−9 to 10−7m) increased [Ca2+]i in fura-2-loaded cells. hUII also caused a rapid and transient activation of ERK1/2 and induction of c-myc. LLCPK1 cells expressed UII mRNA and its receptor GPR14 mRNA, as determined by RT-PCR, and released UII-like immunoreactivity into media. Neutralization of endogenous UII by anti-hUII antibody, but not nonimmune serum, significantly suppressed DNA synthesis. These data suggest that hUII is an autocrine/paracrine growth factor for renal epithelial cells via activation of both protein kinase C and ERK1/2 pathways as well as Ca2+ influx via voltage-dependent Ca2+ channels.

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

1992 ◽  
Vol 284 (3) ◽  
pp. 725-732 ◽  
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.

The macrophage capacity for phagocytosis

1992 ◽  
Vol 101 (4) ◽  
pp. 907-913 ◽  
Author(s):  
G.J. Cannon ◽  
J.A. Swanson
Keyword(s):  
Surface Area ◽  
Membrane Surface ◽  
Fc Receptors ◽  
Phagocytic Capacity ◽  
Membrane Surface Area ◽  
Murine Bone Marrow ◽  
Latex Beads ◽  
Endocytic Compartments ◽  
Frustrated Phagocytosis

Murine bone marrow-derived macrophages, which measure 13.8 +/− 2.3 microns diameter in suspension, can ingest IgG-opsonized latex beads greater than 20 microns diameter. A precise assay has allowed the determination of the phagocytic capacity, and of physiological parameters that limit that capacity. Ingestion of beads larger than 15 microns diameter required IgG-opsonization, and took 30 minutes to reach completion. Despite the dependence on Fc-receptors for phagocytosis of larger beads, cells reached their limit before all cell surface Fc-receptors were occupied. The maximal membrane surface area after frustrated phagocytosis of opsonized coverslips was similar to the membrane surface area required to engulf particles at the limiting diameter, indicating that the capacity was independent of particle shape. Vacuolation of the lysosomal compartment with sucrose, which expanded endocytic compartments, lowered the phagocytic capacity. This decrease was reversed when sucrose vacuoles were collapsed by incubation of cells with invertase. These experiments indicate that the phagocytic capacity is limited by the amount of available membrane, rather than by the availability of Fc-receptors. The capacity was also reduced by depolymerization of cytoplasmic microtubules with nocodazole. Nocodazole did not affect the area of maximal cell spreading during frustrated phagocytosis, but did alter the shape of the spread cells. Thus, microtubules may coordinate cytoplasm for engulfment of the largest particles.

scholarly journals Effect Of Surface Area Of Dialyzer Membrane On The Adequacy Of Haemodialysis

2012 ◽  
Vol 7 (2) ◽  
pp. 9-11 ◽  
Author(s):  
NS Chowdhury ◽  
FMM Islam ◽  
F Zafreen ◽  
BA Begum ◽  
N Sultana ◽  
...  
Keyword(s):  
Renal Disease ◽  
Surface Area ◽  
End Stage Renal Disease ◽  
Membrane Surface ◽  
Reduction Ratio ◽  
Urea Clearance ◽  
Membrane Surface Area ◽  
Stage Renal Disease ◽  
End Stage ◽  
Urea Reduction Ratio

Introduction: Patients with end stage renal disease require 12 hours of haemodialysis per week in three equal sessions (4 hours/day for 3 days/week). But the duration and frequency of treatment can be reduced by increasing the surface area of the dialyzer membrane. Methods: In this prospective study 40 patients of end stage renal disease receiving haemodialysis for more than six months were included to observe the effects of increment in the surface area of the dialyzer membrane on the adequacy of haemodialysis. Result: It was observed that 20 patients receiving haemodialysis on a dialyzer with membrane surface area of 1.2 m² did not have satisfactory solute clearance index. Urea reduction ratio was 45.9 ± 3.03 and fractional urea clearance (Kt/V) was 0.76 ± 0.09. On the other hand patients (20 cases) receiving haemodialysis on a dialyzer with membrane surface area of 1.3 m² had a urea reduction ratio 50.76± 5.16 and fractional urea clearance (Kt/V) 0.91 ± 0.16. All the patients of both groups received dialysis for 8 hours/week in two equal sessions (4 hours/day for 2 days/week). Statistically the increment was significant (p<0.001). Conclusion: This study reveals, adequacy of dialysis can be increased by increasing the surface area of the dialyzer membrane. So, considering the poor socioeconomic condition of Bangladesh and patients' convenience, a short duration, low cost dialysis regime can be tried by increasing the surface area of dialyzer membrane. DOI: http://dx.doi.org/10.3329/jafmc.v7i2.10387 JAFMC 2011; 7(2): 9-11

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