scholarly journals High throughput transepithelial electrical resistance (TEER) measurements on perfused membrane-free epithelia

Lab on a Chip ◽  
2021 ◽  
Author(s):  
A. Nicolas ◽  
F. Schavemaker ◽  
K. Kosim ◽  
D. Kurek ◽  
M. Haarmans ◽  
...  
Keyword(s):  
High Throughput ◽  
Electrical Resistance ◽  
Drug Exposure ◽  
Transepithelial Electrical Resistance ◽  
Ease Of Use ◽  
Inflammatory Processes

We present an instrument for simultaneously measuring TEER in up to 80 perfused epithelial tubules on an OrganoPlate. The sensitivity, speed and ease of use enables screening of tubules during formation, drug exposure and inflammatory processes.

scholarly journals Interleukin-4 and Interleukin- 13 Act on Glomerular Visceral Epithelial Cells

2000 ◽  
Vol 11 (3) ◽  
pp. 413-422
Author(s):  
JOSÉ G. VAN DEN BERG ◽  
JAN ATEN ◽  
M. ANWAR CHAND ◽  
NIKE CLAESSEN ◽  
LISETTE DIJKINK ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Epithelial Cells ◽  
Electrical Resistance ◽  
Structural Changes ◽  
Cell Structure ◽  
Short Circuit ◽  
Transepithelial Electrical Resistance ◽  
Interleukin 4 ◽  
Direct Effects ◽  
Direct Role

Abstract. In minimal change nephrosis (MCN), proteinuria is associated with structural changes of the glomerular visceral epithelial cells (GVEC). The occurrence of MCN has been associated with T-helper2 lymphocyte-dependent conditions. To examine a direct role for type 2 cytokines in GVEC injury, the expression of interleukin (IL)-4/IL-13 receptors by GVEC and direct effects of IL-4 and IL-13 on GVEC were studied. Reverse transcription-PCR showed that isolated human and rat glomeruli and cultured human and rat GVEC expressed mRNA for IL-4Rα, IL-13Rα1, and IL-13Rα2. Protein expression of IL-4Rα and IL-13Rα2 by GVEC in human kidney biopsies and by cultured human GVEC was detected by immunohistochemistry. Western blotting demonstrated phosphorylation of STAT6 in cultured GVEC upon incubation with IL-4 or IL-13. This indicated signal transduction via the heterodimeric receptor complex IL-4R2, which is composed of the IL-4Rα and the IL-13Rα1. Direct effects on GVEC function were examined in monolayer experiments. IL-4 and IL-13 dose-dependently decreased transepithelial electrical resistance of monolayers of rat GVEC to approximately 30 and 40% of baseline values, respectively. The transepithelial electrical resistance decrease was associated with a significant increase in short-circuit current, whereas no changes were observed in the transmonolayer flux of the macromolecules horseradish peroxidase (molecular weight, 44 kD) and 14C-mannitol (molecular weight, 182 Da). No changes in cell structure were observed with electron microscopy. It is concluded that by binding to specific IL-4/IL-13 receptors, IL-4 and IL-13 can exert specific effects on GVEC function, which could be of pathogenetic relevance for glomerular injury in MCN.

Transepithelial Resistance and Inulin Permeability as Endpoints in In Vitro Nephrotoxicity Testing

2002 ◽  
Vol 30 (2_suppl) ◽  
pp. 53-59 ◽  
Author(s):  
Tracey Duff ◽  
Simon Carter ◽  
Gemma Feldman ◽  
Gordon McEwan ◽  
Walter Pfaller ◽  
...  
Keyword(s):  
Proximal Tubule ◽  
Electrical Resistance ◽  
Mdck Cells ◽  
Ussing Chamber ◽  
Fluorescein Isothiocyanate ◽  
Transepithelial Electrical Resistance ◽  
Madin Darby Canine Kidney ◽  
Darby Canine Kidney ◽  
Canine Kidney

Transepithelial electrical resistance (RT) and the flux of fluorescein isothiocyanate (FITC) across Madin Darby canine kidney (MDCK) strain 1 cells and porcine epithelial kidney (LLC-PK1) monolayers were compared between three laboratories for a range of nephrotoxins. The precision of the REMS AutoSampler was similar to that of the Ussing chamber and the ENDOHM® technique, but superior to using chopstick electrodes, for measurements of resistance. The nephrotoxins used were selective for the proximal tubule, and in all cases, LLC-PK1 cells were more sensitive than MDCK cells. In most cases, change in RT was a more sensitive indicator of damage than alterations in FITC flux. The REMS system provides high intra-plate precision for RT measurements and is a higher throughput system, which is applicable to screening for nephrotoxicity in vitro.

Transport properties of toad kidney epithelia in culture

1981 ◽  
Vol 241 (3) ◽  
pp. C154-C159 ◽  
Author(s):  
F. M. Perkins ◽  
J. S. Handler
Keyword(s):  
Electrical Resistance ◽  
Hormonal Regulation ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Transepithelial Electrical Resistance ◽  
Apical Surface ◽  
Osmotic Water ◽  
Sodium Flux ◽  
Regulation Of Transport ◽  
Osmotic Water Flow

The characteristics of a continuous line of toad kidney epithelial cells (A6) are described. These cells form a monolayer epithelium of high transepithelial electrical resistance (about 5,000 omega . cm2). The cells generate a transepithelial potential difference (apical surface negative) of about 9 mV. The short-circuit current is equivalent to net sodium flux. Net sodium flux is stimulated by aldosterone and by analogues of cAMP. The stimulation is readily reversible. Neither urea permeability nor osmotic water flow is altered by analogues of cAMP. Amiloride eliminates 90% of the short-circuit current. Thus A6 cells form an epithelium with several differentiated properties including hormonal regulation of transport.

scholarly journals 230 Genome editing applications in plants: high-throughput CRISPR/Cas editing for crop improvement

2019 ◽  
Vol 97 (Supplement_3) ◽  
pp. 56-56
Author(s):  
Michael Thomson
Keyword(s):  
Genome Editing ◽  
High Throughput ◽  
Positional Cloning ◽  
Gene Editing ◽  
Crop Improvement ◽  
Cost Effective ◽  
Gene Families ◽  
Ease Of Use ◽  
Plant Genome ◽  
Wide Range

Abstract The precision and ease of use of CRISPR nucleases, such as Cas9 and Cpf1, for plant genome editing has the potential to accelerate a wide range of applications for crop improvement. For upstream research on gene discovery and validation, rapid gene knock-outs can enable testing of single genes and multi-gene families for functional effects. Large chromosomal deletions can test the function of tandem gene arrays and assist with positional cloning of QTLs by helping to narrow down the target region. Nuclease-deactivated Cas9 fusion proteins with transcriptional activators and repressors can be used to up and down-regulate gene expression. Even more promising, gene insertions and allele replacements can provide the opportunity to rapidly test the effects of different alleles at key loci in the same genetic background, providing a more precise alternative to marker-assisted backcrossing. Recently, Texas A&M AgriLife Research has supported the development of a Crop Genome Editing Lab at Texas A&M working towards optimizing a high-throughput gene editing pipeline and providing an efficient and cost-effective gene editing service for research and breeding groups. The lab is using rice as a model to test and optimize new approaches aimed towards overcoming current bottlenecks. For example, a wealth of genomics data from the rice community enables the development of novel approaches to predict which genes and target modifications may be most beneficial for crop improvement, taking advantage of known major genes, high-resolution GWAS data, multiple high-quality reference genomes, transcriptomics data, and resequencing data from the 3,000 Rice Genomes Project. Current projects have now expanded to work across multiple crops to provide breeding and research groups with a rapid gene editing pipeline to test candidate genes in their programs, with the ultimate goal of developing nutritious, high-yielding, stress-tolerant crops for the future.

Na+-sugar cotransport system as a polarization marker during organization of epithelial membrane

1988 ◽  
Vol 255 (6) ◽  
pp. C745-C753 ◽  
Author(s):  
C. Lasheras ◽  
J. A. Scott ◽  
C. A. Rabito
Keyword(s):  
Electrical Resistance ◽  
Apical Membrane ◽  
Sugar Transport ◽  
Cell Polarization ◽  
Transepithelial Electrical Resistance ◽  
Apical Surface ◽  
Binding Studies ◽  
Occluding Junctions ◽  
Basolateral Side ◽  
Epithelial Membranes

The present study analyzed the changes in Na+-dependent sugar transport and transepithelial electrical resistance as LLC-PK1 cells reorganize into epithelial membranes. Sugar influx increased to reach a maximum 9 h after plating. The increase in the transepithelial electrical resistance, however, showed a significant delay, reaching steady state 15 h after plating. No changes in the electrochemical Na+ gradient were observed during the reorganization of the epithelial membranes. Kinetic analysis and [3H]phlorizin-binding studies showed that the increase in sugar influx resulted from an increase in the number of carriers. Unidirectional sugar influx measurements indicated that the sugar transporters were primarily located at the apical surface of the epithelial cells. These observations are consistent with the hypothesis that the sorting of native proteins occurs intracellularly before their insertion in the apical membrane, or as an alternative that they are randomly inserted, but then immediately sorted such as any carrier could be detected in the basolateral side during the reorganization process. In addition, the results suggest that the functional development of the apical membrane may occur before the complete sealing of the intercellular space during the development of the occluding junctions. Furthermore, development of the sugar transport system and occluding junctions was inhibited by cycloheximide and puromycin but not by actinomycin D, suggesting that the expression of epithelial cell polarization is probably a posttranslational event in the protein synthesis.

scholarly journals DNA metabarcoding of insects and allies: an evaluation of primers and pipelines

2015 ◽  
Vol 105 (6) ◽  
pp. 717-727 ◽  
Author(s):  
G.-J. Brandon-Mong ◽  
H.-M. Gan ◽  
K.-W. Sing ◽  
P.-S. Lee ◽  
P.-E. Lim ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Illumina Miseq ◽  
Ease Of Use ◽  
Malaise Trap ◽  
Detection Rates ◽  
Operational Taxonomic Units ◽  
Arthropod Species ◽  
Primer Sets ◽  
Arthropod Biodiversity

AbstractMetabarcoding, the coupling of DNA-based species identification and high-throughput sequencing, offers enormous promise for arthropod biodiversity studies but factors such as cost, speed and ease-of-use of bioinformatic pipelines, crucial for making the leapt from demonstration studies to a real-world application, have not yet been adequately addressed. Here, four published and one newly designed primer sets were tested across a diverse set of 80 arthropod species, representing 11 orders, to establish optimal protocols for Illumina-based metabarcoding of tropical Malaise trap samples. Two primer sets which showed the highest amplification success with individual specimen polymerase chain reaction (PCR, 98%) were used for bulk PCR and Illumina MiSeq sequencing. The sequencing outputs were subjected to both manual and simple metagenomics quality control and filtering pipelines. We obtained acceptable detection rates after bulk PCR and high-throughput sequencing (80–90% of input species) but analyses were complicated by putative heteroplasmic sequences and contamination. The manual pipeline produced similar or better outputs to the simple metagenomics pipeline (1.4 compared with 0.5 expected:unexpected Operational Taxonomic Units). Our study suggests that metabarcoding is slowly becoming as cheap, fast and easy as conventional DNA barcoding, and that Malaise trap metabarcoding may soon fulfill its potential, providing a thermometer for biodiversity.

scholarly journals The effect of neutrophil migration on epithelial permeability.

1986 ◽  
Vol 103 (6) ◽  
pp. 2729-2738 ◽  
Author(s):  
L C Milks ◽  
G P Conyers ◽  
E B Cramer
Keyword(s):  
Electrical Resistance ◽  
Neutrophil Migration ◽  
Inflammatory Lesion ◽  
Time Frame ◽  
Transepithelial Electrical Resistance ◽  
Human Neutrophils ◽  
Epithelial Permeability ◽  
Vitro Model ◽  
Migratory Cells

To reach an inflammatory lesion, neutrophils must frequently traverse the epithelium of an infected organ. Whether the actual migration of neutrophils alters the epithelial permeability is unknown. Through the use of an in vitro model system it was possible to directly determine the effect of neutrophil emigration on the transepithelial electrical resistance of the monolayer. Human neutrophils (5 X 10(6) cells/ml) were placed in the upper compartment of a combined chemotaxis/resistance chamber and stimulated for 40 min by a gradient of 10(-7) M n-formyl-methionyl-leucyl-phenylalanine to traverse a confluent monolayer of canine kidney epithelial cells grown on micropore filters. Neither the chemoattractant alone (10(-5)-10(-9) M) nor the accumulation of an average of eight neutrophils per millimeter of epithelium lowered the transepithelial electrical resistance. However, under certain conditions the migration of neutrophils temporarily increased the permeability of the monolayer. The resistance fell approximately 48% within 5 min if the migratory cells were stimulated to reverse their migration across the same monolayer. As re-migration continued, the resistance returned to its initial levels within 60 min. Doubling the initial neutrophil concentration to 10 X 10(6) cells/ml resulted in the accumulation of an average of 66 neutrophils per millimeter of epithelium and an average fall in resistance of 46% (r = 0.98; P less than 0.001) in 40 min. If the resistance had fallen less than 45%, removal of the neutrophils remaining in the upper compartment resulted in a return of the transepithelial electrical resistance to its initial level within 65 min. However, when the fall was greater than 45%, the resistance only recovered to 23.5% of its initial levels within the same time frame. Thus, these results suggest that the integrity of an epithelium can, under certain conditions, be affected by the emigration of neutrophils, but that this effect is either completely or partially reversible within 65 min.

Online monitoring of transepithelial electrical resistance (TEER) in an apparatus for combined dissolution and permeation testing

2010 ◽  
Vol 392 (1-2) ◽  
pp. 134-140 ◽  
Author(s):  
Marco Muendoerfer ◽  
Ulrich F. Schaefer ◽  
Petra Koenig ◽  
Jutta S. Walk ◽  
Petra Loos ◽  
...  
Keyword(s):  
Electrical Resistance ◽  
Online Monitoring ◽  
Transepithelial Electrical Resistance
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