Case 8: Status of the of P-gp-Mediated Transport Through Confluent Cell Monolayers

2021 ◽  
pp. 737-763
Author(s):  
Joe Bentz ◽  
Harma Ellens
Keyword(s):  
Cell Monolayers ◽  
Confluent Cell

scholarly journals Dense active matter model of motion patterns in confluent cell monolayers

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Silke Henkes ◽  
Kaja Kostanjevec ◽  
J. Martin Collinson ◽  
Rastko Sknepnek ◽  
Eric Bertin
Keyword(s):  
Active Matter ◽  
Motion Patterns ◽  
Cell Monolayers ◽  
Matter Model ◽  
Confluent Cell

scholarly journals Cell–cell Interaction Underlies Formation of Fluid in the Male Reproductive Tract of the Rat

2005 ◽  
Vol 125 (5) ◽  
pp. 443-454 ◽  
Author(s):  
King-ho Cheung ◽  
George P.H. Leung ◽  
Matthew C.T. Leung ◽  
Winnie W.C. Shum ◽  
Wen-liang Zhou ◽  
...  
Keyword(s):  
Reproductive Tract ◽  
Cell Types ◽  
Cell Interaction ◽  
Fluorescence Measurement ◽  
Basal Cells ◽  
Anion Secretion ◽  
Cell Monolayers ◽  
Principal Cells ◽  
Whole Cell Patch Clamp ◽  
Confluent Cell

The epithelia lining the epididymides of many species consists of several cell types. We have provided evidence that the basal cells are essential to the integrated functions of the epithelium. Basal cells, but not principal cells, and other cells in the epididymis express TRPC3 and COX-1. We have isolated basal cells from intact rat epididymis using antibody-coated Dynabeads and subjected them to whole-cell patch-clamp measurement of nonselective cation channel activity, a feature of TRPC3 protein, and Fluo-3 fluorescence measurement of intracellular Ca2+ concentration. The results show that a nonselective cation current blockable by La3+ (0.1 mM), Gd3+ (0.1 mM), or SKF96365 (20 μM) could be activated by lysylbradykinin (200 nM). In cells loaded with Fluo-3, addition of lysylbradykinin (100 nM) caused a sustained increase of intracellular Ca2+. This effect was blocked by Gd3+ (0.1 mM) or SKF96365 (20 μM) and was not observed in Fluo-3–loaded principal cells. Stimulation of basal cell/principal cell cocultures with lysylbradykinin (200 nM) evoked in principal cells a current with CFTR-Cl− channel characteristics. Isolated principal cells in the absence of basal cells did not respond to lysylbradykinin but responded to PGE2 (100 nM) with activation of a CFTR-like current. Basal cells, but not principal cells, released prostaglandin E2 when stimulated with lysylbradykinin (100 nM). The release was blocked by SKF96365 (20 μM) and BAPTA-AM (0.05 or 0.1 mM). Confluent cell monolayers harvested from a mixture of disaggregated principal cells and basal cells responded to lysylbradykinin (100 nM) and PGE2 (500 nM) with an increase in electrogenic anion secretion. The former response was dependent on prostaglandin synthesis as piroxicam blocked the response. However, cell cultures obtained from principal cells alone responded to PGE2 but not to bradykinin. These results support the notion that basal cells regulate principal cells through a Ca2+ and COX signaling pathway.

scholarly journals Expression of epithelial Na channels in Xenopus oocytes.

1990 ◽  
Vol 96 (1) ◽  
pp. 23-46 ◽  
Author(s):  
L G Palmer ◽  
I Corthesy-Theulaz ◽  
H P Gaeggeler ◽  
J P Kraehenbuhl ◽  
B Rossier
Keyword(s):  
Sedimentation Coefficient ◽  
High Sensitivity ◽  
Sucrose Density Gradient ◽  
Na Channel ◽  
Channel Activity ◽  
Membrane Expression ◽  
Cell Monolayers ◽  
A6 Cells ◽  
Epithelial Na Channels ◽  
Confluent Cell

Epithelial Na channel activity was expressed in oocytes from Xenopus laevis after injection of mRNA from A6 cells, derived from Xenopus kidney. Poly A(+) RNA was extracted from confluent cell monolayers grown on either plastic or permeable supports. 1-50 ng RNA was injected into stage 5-6 oocytes. Na channel activity was assayed as amiloride-sensitive current (INa) under voltage-clamp conditions 1-3 d after injection. INa was not detectable in noninjected or water-injected oocytes. This amiloride-sensitive pathway induced by the mRNA had a number of characteristics in common with that in epithelial cells, including (a) high selectivity for Na over K, (b) high sensitivity to amiloride with an apparent K1 of approximately 100 nM, (c) saturation with respect to external Na with an apparent Km of approximately 10 mM, and (d) a time-dependent activation of current with hyperpolarization of the oocyte membrane. Expression of channel activity was temperature dependent, being slow at 19 degrees C but much more rapid at 25 degrees C. Fractionation of mRNA on a sucrose density gradient revealed that the species of RNA inducing channel activity had a sedimentation coefficient of approximately 17 S. Treatment of filter-grown cells with 300 nM aldosterone for 24 h increased Na transport in the A6 cells by up to fivefold but did not increase the ability of mRNA isolated from those cells to induce channel activity in oocytes. The apparent abundance of mRNA coding for channel activity was 10-fold less in cells grown on plastic than in those grown on filters, but was increased two- to threefold by aldosterone.

scholarly journals Temperature-responsive intelligent interfaces for biomolecular separation and cell sheet engineering

2009 ◽  
Vol 6 (suppl_3) ◽  
Author(s):  
Kenichi Nagase ◽  
Jun Kobayashi ◽  
Teruo Okano
Keyword(s):  
Cell Sheet ◽  
Copolymer Composition ◽  
Cell Sheets ◽  
Chromatographic Separations ◽  
Cell Sheet Engineering ◽  
External Temperature ◽  
Cell Monolayers ◽  
Temperature Responsive ◽  
Confluent Cell ◽  
Dependent Cell

Temperature-responsive intelligent surfaces, prepared by the modification of an interface with poly( N -isopropylacrylamide) and its derivatives, have been used for biomedical applications. Such surfaces exhibit temperature-responsive hydrophilic/hydrophobic alterations with external temperature changes, which, in turn, result in thermally modulated interactions with biomolecules and cells. In this review, we focus on the application of these intelligent surfaces to chromatographic separation and cell cultures. Chromatographic separations using several types of intelligent surfaces are mentioned briefly, and various effects related to the separation of bioactive compounds are discussed, including wettability, copolymer composition and graft polymer architecture. Similarly, we also summarize temperature-responsive cell culture substrates that allow the recovery of confluent cell monolayers as contiguous living cell sheets for tissue-engineering applications. The key factors in temperature-dependent cell adhesion/detachment control are discussed from the viewpoint of grafting temperature-responsive polymers, and new methodologies for effective cell sheet culturing and the construction of thick tissues are summarized.

Effect of high extracellular K+ on Na-K-ATPase in cultured canine kidney cells

1990 ◽  
Vol 259 (2) ◽  
pp. F227-F232 ◽  
Author(s):  
M. A. Manuli ◽  
I. S. Edelman
Keyword(s):  
Atpase Activity ◽  
Northern Analysis ◽  
Beta Subunits ◽  
Cell Monolayers ◽  
High K ◽  
Confluent Cell ◽  
K Transport ◽  
Darby Canine Kidney ◽  
Canine Kidney ◽  
Specific Mrna

The Madin-Darby canine kidney (MDCK) cell line was used to evaluate the influence of high extracellular K+, independent of hormonal effects, on renal Na-K-adenosinetriphosphatase (ATPase) activity and abundance. Confluent cell monolayers were incubated in control (5 mM) or high K+ (7.5 mM) medium for 24 h. Exposure to high K+ elicited a 46% rise in Na-K-ATPase activity and a 55% increase in ouabain-sensitive 86Rb uptake. Na-K-ATPase abundance, estimated from the number of ouabain-binding sites, also increased 63% over control in cells exposed to 7.5 mM K+, and as a consequence there was no statistically significant change in the catalytic turnover number. Northern blot analysis using rat cDNA probes for the alpha 1- and beta-subunits showed no corresponding changes in subunit-specific mRNA abundances at 24 h. We conclude that chronic exposure to high extracellular K+ produces a rise in renal epithelial Na-K-ATPase activity and active K+ transport, independent of changes in aldosterone, renal blood flow, or extracellular Na+ concentration. This effect is due to an increase in enzyme abundance rather than a change in catalytic turnover rate. The results of Northern analysis suggest that regulation of Na-K-ATPase activity and abundance by high K+ may involve translational or posttranslational mechanisms, but further study with cDNA probes of canine origin is needed to resolve this issue.

A chip device to determine surface charge properties of confluent cell monolayers by measuring streaming potential

Lab on a Chip ◽  
2020 ◽  
Vol 20 (20) ◽  
pp. 3792-3805 ◽  
Author(s):  
András Kincses ◽  
Ana R. Santa-Maria ◽  
Fruzsina R. Walter ◽  
László Dér ◽  
Nóra Horányi ◽  
...  
Keyword(s):  
Cell Surface ◽  
Surface Charge ◽  
Streaming Potential ◽  
Cell Monolayers ◽  
Biological Barriers ◽  
Cell Surface Charge ◽  
Cell Layers ◽  
Confluent Cell

Chip device to monitor streaming potential of confluent cell layers reflecting cell surface charge important for the function of biological barriers.

Role of connexins in microvascular dysfunction during inflammation

2011 ◽  
Vol 89 (1) ◽  
pp. 1-12 ◽  
Author(s):  
Karel Tyml
Keyword(s):  
Cultured Cells ◽  
Electrical Coupling ◽  
Microvascular Dysfunction ◽  
Microvascular Endothelial Cells ◽  
Dependent Manner ◽  
Cell Monolayers ◽  
Confluent Cell ◽  
Diameter Change

In arterioles, a locally initiated diameter change can propagate rapidly along the vessel length (arteriolar conducted response), thus contributing to arteriolar hemodynamic resistance. The response is underpinned by electrical coupling along the arteriolar endothelial layer. Connexins (Cx; constituents of gap junctions) are required for this coupling. This review addresses the effect of acute systemic inflammation (sepsis) on arteriolar conduction and interendothelial electrical coupling. Lipopolysaccharide (LPS; an initiating factor in sepsis) and polymicrobial sepsis (24 h model) attenuate conducted vasoconstriction in mice. In cultured microvascular endothelial cells harvested from rat and mouse skeletal muscle, LPS reduces both conducted hyperpolarization–depolarization along capillary-like structures and electrical coupling along confluent cell monolayers. LPS also tyrosine-phosphorylates Cx43 and serine-dephosphorylates Cx40. Since LPS-reduced coupling is Cx40- but not Cx43-dependent, only Cx40 dephosphorylation may be consequential. Nitric oxide (NO) overproduction is critical in advanced sepsis, since the removal of this overproduction prevents the attenuated conduction. Consistently, (i) exogenous NO in cultured cells reduces coupling in a Cx37-dependent manner, and (ii) the septic microvasculature in vivo shows no Cx40 phenotype. A complex role emerges for endothelial connexins in sepsis. Initially, LPS may reduce interendothelial coupling and arteriolar conduction by targeting Cx40, whereas NO overproduction in advanced sepsis reduces coupling and conduction by targeting Cx37 instead.

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