Improved Trans-endothelial Electrical Resistance Sensing using Microfluidic Low-Temperature Co-fired Ceramics

2013 ◽  
Vol 2013 (CICMT) ◽  
pp. 000162-000167
Author(s):  
William L. Mercke ◽  
Thomas Dziubla ◽  
Richard E. Eitel ◽  
Kimberly Anderson
Keyword(s):  
Endothelial Cell ◽  
Low Temperature ◽  
Electrical Resistance ◽  
Cell Monolayer ◽  
Three Dimensional ◽  
Barrier Properties ◽  
Ceramic Materials ◽  
Functional Change ◽  
Cell Monolayers ◽  
Lock In

Trans-endothelial Electrical Resistance (TEER) and cellular impedance measurements are widely used to evaluate the barrier properties and functional change of endothelial cell monolayers. In the current work, low temperature cofired ceramics (LTCC) are applied enabling the incorporation of TEER and impediametric measurements in an integrated microfluidic chip. LTCC materials are an ideal substrate for biomedical and cell-based microfluidics due to their biocompatibility and ability to combine complex three dimensional structures with optical, fluidic, and electrical functionality. Multilayer microfluidic ceramic devices incorporating gold measurement electrodes where prepared using standard LTCC manufacturing procedures. The sensitivity of the resulting LTCC devices were compared to systems currently on the market for TEER measurements. These results indicate the LTCC device is able to effectively detect the growth of an endothelial cell monolayer. Results further evaluate endothelial cell viability using electrical resistance and Live/Dead assay. Finally, the results from this study also display improved sensitivity through the optimization of the electrode geometry and use of a lock-in amplifier. These results provide a solid basis for using low temperature co-fired ceramic materials for microfluidic TEER devices.

Characterization of an In Vitro Model to Study the Permeability of Human Arterial Endothelial Cell Monolayers

1988 ◽  
Vol 60 (02) ◽  
pp. 240-246 ◽  
Author(s):  
Erna G Langeler ◽  
Victor W M van Hinsbergh
Keyword(s):  
Endothelial Cell ◽  
Calcium Ions ◽  
Electrical Resistance ◽  
Serum Proteins ◽  
Cell Monolayer ◽  
Extracellular Calcium ◽  
Endothelial Cell Monolayer ◽  
Passage Rate ◽  
Cell Monolayers

SummaryA model has been developed to study the transport of fluid and macromolecules through human arterial umbilical cord endothelial cell monolayers in vitro. Cells were cultured on fibronectin- coated polycarbonate filters and formed within a few days a tight monolayer, with an electrical resistance of 17 ± 4 Ohm · cm2. The cells were connected by close cell contacts with tight junctions. The passáge-rate of horse radish peroxidase (HRP) through these filters was 20-40 fold lower than through filters without an endothelial monolayer. The continuous presence of 10% human serum was needed to maintain the electrical resistance of the monolayer and its barrier function towards macromolecules. Chelation of extracellular calcium resulted in an increased permeability and a decreased electrical resistance of the monolayer. This process was reversible by re-addition of calcium ions to the cells. The permeation rate of dextrans of various molecular weights (9-480 kD) was inversely related to the molecular mass of the molecule. No difference was measured between the passage rate of dextran of 480 kD and dextran of 2,000 kD. Incubation of the endothelial cell monolayer with 2-deoxy-D-glucose resulted in a decreased permeability but it had no effect on electrical resistance. This suggests that the passage-process is energy- dependent.Fluid permeation through the endothelial cell monolayer on filters was measured in a perfusion chamber under 20 mmHg hydrostatic pressure. It was decreased by the presence of serum proteins and responded reversibly on the chelation and readdition of extracellular calcium ions.

scholarly journals Effects of 1,2-Dimethylhydrazine on Barrier Properties of Rat Large Intestine and IPEC-J2 Cells

2021 ◽  
Vol 22 (19) ◽  
pp. 10278
Author(s):  
Viktoria Bekusova ◽  
Linda Droessler ◽  
Salah Amasheh ◽  
Alexander G. Markov
Keyword(s):  
Colorectal Cancer ◽  
Electrical Resistance ◽  
Barrier Properties ◽  
Acute Effect ◽  
Paracellular Permeability ◽  
Transepithelial Electrical Resistance ◽  
Rat Colon ◽  
Sodium Fluorescein ◽  
Cell Monolayers

Colon cancer is accompanied by a decrease of epithelial barrier properties, which are determined by tight junction (TJ) proteins between adjacent epithelial cells. The aim of the current study was to analyze the expression of TJ proteins in a rat model of 1,2-dimethylhydrazine (DMH)-induced colorectal cancer, as well as the barrier properties and TJ protein expression of IPEC-J2 cell monolayers after incubation with DMH. Transepithelial electrical resistance and paracellular permeability for sodium fluorescein of IPEC-J2 were examined by an epithelial volt/ohm meter and spectrophotometry. The expression and localization of TJ proteins were analyzed by immunoblotting and immunohistochemistry. In the colonic tumors of rats with DMH-induced carcinogenesis, the expression of claudin-3 and -4 was significantly increased compared to controls. The transepithelial electrical resistance of IPEC-J2 cells increased, while paracellular permeability for sodium fluorescein decreased, accompanied by an increased expression of claudin-4. The increase of claudin-4 in rat colon after chronic DMH exposure was consistent with the acute effect of DMH on IPEC-J2 cells, which may indicate an essential role of this protein in colorectal cancer development.

THE BINDING OF ANTITHROMBIN TO CAPILLARY ENDOTHELIAL CELLS GROWN IN VITRO

1987 ◽  
Author(s):  
A de Agostini ◽  
J Marcum ◽  
R Rosenberg
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Heparan Sulfate ◽  
Cell Monolayer ◽  
Whole Cells ◽  
Cell Monolayers ◽  
Bacterial Enzyme ◽  
Capillary Endothelial Cells ◽  
Binding Cell

Cloned endothelial cells from rat epididymal fat pads synthesize anticoagulantly active heparan sulfate proteoglycans containing the disaccharide, GlcA→ AMN-3,6-O-SO3, which is a marker for the antithrombin-binding domain of heparin. To demonstrate that antithrombin (AT) binds to cell surface heparan sulfate, a binding assay employing 125I-AT and cell monolayers has been developed. Post-confluent endothelial cells (7 days) were incubated with radiolabeled AT for 1 h at 4° and washed with PBS. Bound radioactivity was quantitated after solubilizing whole cells. Under these conditions, ∼1% (2174±50 cpm/5x104 cells) of the 125I-AT bound to the endothelial cell monolayer, whereas none of the radiolabeled protein bound to CHO cells or bovine smooth muscle cells. Utilization of unlabeled AT (1 μM) in experiments conducted as described above resulted in a reduction (73%) of the binding of the labeled species to endothelial cells. To assess whether heparan sulfate was responsible for AT binding, cell monolayers were incubated for 1 h at 37° with purified Flavobacterium heparinase (0.2 units). Over 90% of 125I-AT binding to these cellular elements was suppressed with the bacterial enzyme. Internalization of radiolabeled AT by endothelial cells was examined by incubating the protease inhibitor and cells at 4° and 37 . An initial rapid binding was observed at both temperatures. At 4° AT binding plateaued within 15 min, whereas at 37° binding did not plateau until 60 min and was 30% greater than that observed at 4. These data suggest that surface-associated AT can be internalized by endothelial cells. In addition, AT binding was shown to increase with the length of endothelial cell postconfluence, indicating an accumulation of heparan sulfate by these cells during quiescence. In conclusion, our studies support the hypothesis that the vascular endothelium is coated with heparan sulfate-bound AT, which is responsible for the antithrombotic properties of these natural surfaces.

scholarly journals Drug Flux Across RPE Cell Models: The Hunt for An Appropriate Outer Blood–Retinal Barrier Model for Use in Early Drug Discovery

Pharmaceutics ◽  
2020 ◽  
Vol 12 (2) ◽  
pp. 176 ◽  
Author(s):  
Laura Hellinen ◽  
Heidi Hongisto ◽  
Eva Ramsay ◽  
Kai Kaarniranta ◽  
Kati-Sisko Vellonen ◽  
...  
Keyword(s):  
Drug Discovery ◽  
Retinal Pigment ◽  
Cell Monolayer ◽  
Barrier Properties ◽  
Cell Models ◽  
Blood Retinal Barrier ◽  
Rpe Cells ◽  
Cell Monolayers ◽  
Value Range ◽  
Drug Flux

The retinal pigment epithelial (RPE) cell monolayer forms the outer blood–retinal barrier and has a crucial role in ocular pharmacokinetics. Although several RPE cell models are available, there have been no systematic comparisons of their barrier properties with respect to drug permeability. We compared the barrier properties of several RPE secondary cell lines (ARPE19, ARPE19mel, and LEPI) and both primary (hfRPE) and stem-cell derived RPE (hESC-RPE) cells by investigating the permeability of nine drugs (aztreonam, ciprofloxacin, dexamethasone, fluconazole, ganciclovir, ketorolac, methotrexate, voriconazole, and quinidine) across cell monolayers. ARPE19, ARPE19mel, and hfRPE cells displayed a narrow Papp value range, with relatively high permeation rates (5.2–26 × 10−6 cm/s. In contrast, hESC-RPE and LEPI cells efficiently restricted the drug flux, and displayed even lower Papp values than those reported for bovine RPE-choroid, with the range of 0.4–32 cm−6/s (hESC-RPE cells) and 0.4–29 × 10−6 cm/s, (LEPI cells). Therefore, ARPE19, ARPE19mel, and hfRPE cells failed to form a tight barrier, whereas hESC-RPE and LEPI cells restricted the drug flux to a similar extent as bovine RPE-choroid. Therefore, LEPI and hESC-RPE cells are valuable tools in ocular drug discovery.

Biocompatibility Evaluation of Human Umbilical Vein Endothelial Cells Directly onto Low-Temperature Co-fired Ceramic Materials for Microfluidic Applications

2012 ◽  
Vol 2012 (CICMT) ◽  
pp. 000549-000556 ◽  
Author(s):  
William L. Mercke ◽  
Thomas Dziubla ◽  
Richard E. Eitel ◽  
Kimberly Anderson
Keyword(s):  
Endothelial Cells ◽  
Low Temperature ◽  
Cellular Response ◽  
Umbilical Vein ◽  
Three Dimensional ◽  
Ceramic Materials ◽  
Human Umbilical Vein ◽  
Electronic Circuitry ◽  
Cellular Attachment ◽  
Umbilical Vein Endothelial Cells

Expansion of Low-Temperature Co-fired Ceramic materials into microfluidic systems technology has many beneficial applications due to their ability to combine complex three dimensional structures with optical, fluidic, electrical functions. Evaluations of the biocompatibility of these Low-Temperature Co-fired Ceramic materials are vital for expanding into biomedical research. The few biocompatibility studies on Low-Temperature Co-fired Ceramics generally show negative cellular response to thick film pastes used in generating the electronic circuitry patterns. In this study, biocompatibility of Human Umbilical Vein Endothelial Cells was examined on Heraeus's Low-Temperature Co-fired Ceramic tape and two of their conductive pastes. The biocompatibility was assessed by monitoring cellular attachment and viability up to three days. This study examines the idea of leachates being detrimental to cells due to a study that suggests the possibility of harmful leachates. Results indicate difficulty in initial attachment of Human Umbilical Vein Endothelial Cells to sintered Low-Temperature Co-fired Ceramic tapes, but no hindrance of cellular attachment and growth onto the two conductive pastes. Outcomes also demonstrate that possible harmful leachates from Low-Temperature Co-fired Ceramic materials don't thwart cellular attachment and growth for up to three days of cell culturing. These results provide a basis for biological devices using Low-Temperature Co-fired Ceramic materials.

scholarly journals Uric acid attenuates trophoblast invasion and integration into endothelial cell monolayers

2009 ◽  
Vol 297 (2) ◽  
pp. C440-C450 ◽  
Author(s):  
Shannon A. Bainbridge ◽  
James M. Roberts ◽  
Frauke von Versen-Höynck ◽  
Jessa Koch ◽  
Lia Edmunds ◽  
...  
Keyword(s):  
Uric Acid ◽  
Endothelial Cell ◽  
Cell Apoptosis ◽  
Cell Monolayer ◽  
Trophoblast Invasion ◽  
Microvascular Endothelial Cell ◽  
Endothelial Cell Monolayer ◽  
Endothelial Cell Apoptosis ◽  
Trophoblast Cells ◽  
Cell Monolayers

Hyperuricemia develops as early as 10 wk of gestation in women who later develop preeclampsia. At this time the invasive trophoblast cells are actively remodeling the uterine spiral arterioles, integrating into and finally replacing the vascular endothelial lining. In the nonpregnant population uric acid has several pathogenic effects on vascular endothelium. We therefore sought to examine the effects of uric acid (0–7 mg/dl) on trophoblast cell invasion through an extracellular matrix using an in vitro Matrigel invasion assay. We also assessed trophoblast integration into a uterine microvascular endothelial cell monolayer in a trophoblast-endothelial cell coculture model. Additionally, we addressed the importance of redox signaling and trophoblast-induced endothelial cell apoptosis. Uric acid elicited a concentration-dependent attenuation of trophoblast invasion and integration into a uterine microvascular endothelial cell monolayer. The attenuated trophoblast integration appeared to be the result of reduced trophoblast-induced endothelial cell apoptosis, likely through the intracellular antioxidant actions of uric acid. In a test of relevance, pooled serum (5% vol/vol) from preeclamptic women attenuated the ability of trophoblast cells to integrate into the endothelial cell monolayers compared with pooled serum from healthy pregnant controls, and this response was partially rescued when endogenous uric acid was previously removed with uricase. Taken together these data support the hypothesis that elevations in circulating uric acid in preeclamptic women contribute to the pathogenesis of the disorder, in part, through attenuation of normal trophoblast invasion and spiral artery vascular remodeling.

Fluid transport across cultured bovine corneal endothelial cell monolayers

1992 ◽  
Vol 262 (1) ◽  
pp. C98-C103 ◽  
Author(s):  
P. Narula ◽  
M. Xu ◽  
K. Y. Kuang ◽  
R. Akiyama ◽  
J. Fischbarg
Keyword(s):  
Endothelial Cell ◽  
Electrical Resistance ◽  
Corneal Endothelium ◽  
Fluid Transport ◽  
Corneal Endothelial Cell ◽  
Potential Difference ◽  
Cell Monolayers ◽  
Transport Fluid ◽  
Bovine Corneal Endothelial Cell

The mammalian corneal endothelium is known to transport fluid from the stromal compartment to the aqueous humor, thereby maintaining corneal transparency. Corneal endothelial cells have been cultured for some years now, but whether they preserve their in vivo ability to actively transport fluid is not known. We have now grown bovine corneal endothelial cell monolayers (BCECM) on permeable substrates (Transwell) and report that, just like their counterparts in vivo, these cultured cells pump fluid from the basal to the apical compartment and display measurable electrical resistance and potential difference across the monolayer. BCECM were grown on collagen-treated permeable supports using Dulbecco's modified Eagle's medium (DMEM)/20% fetal bovine serum with antibiotics. Cells grew to confluence in 5-7 days and displayed polygonal shape. Only cells from passages 1-3 were utilized. Inserts were fitted directly into Lucite chambers specially built. The rate of fluid pumping by BCECM was 3.96 +/- 0.49 (SE) microliter.h-1.cm-2 (n = 13) and could be measured continuously for several hours; fluid pumping was inhibited by 0.2 mM amiloride. The specific electrical resistance of the monolayers was 180 +/- 22 omega.cm2 (n = 11). A mean electrical potential difference of 63.8 +/- 3.7 microV (n = 15, range 40-100 microV, apical side negative) was recorded across the monolayers in DMEM. The availability of the commercial inserts makes this procedure practical; as a consequence, the rate of fluid transport by cultured corneal endothelium has been quantitated for the first time. This method can now be extended to other cultured layers.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Three-dimensional Monolayer Stress Microscopy

2019 ◽  
Author(s):  
Ricardo Serrano ◽  
Aereas Aung ◽  
Yi-Ting Yeh ◽  
Shyni Varghese ◽  
Juan C. Lasheras ◽  
...  
Keyword(s):  
Flexible Substrate ◽  
Cell Monolayer ◽  
Three Dimensional ◽  
Lateral Deformation ◽  
Equilibrium Equations ◽  
Island Size ◽  
Cell Monolayers ◽  
Bending Stresses ◽  
A Cell ◽  
External Loads

ABSTRACTMany biological processes involve the collective generation and transmission of mechanical stresses across cell monolayers. In these processes, the monolayer undergoes lateral deformation and bending due to the tangential and normal components of the cell-generated stresses. Monolayer Stress Microscopy (MSM) methods have been developed to measure the intracellular stress distribution in cell monolayers. However, current methods assume plane monolayer geometry and neglect the contribution of bending to the intracellular stresses.This work introduces a three-dimensional (3D) MSM method that calculates monolayer stress from measurements of the 3D traction stresses exerted by the cells on a flexible substrate. The calculation is carried out by imposing equilibrium of forces and moments in the monolayer, subject to external loads given by the 3D traction stresses. The equilibrium equations are solved numerically, and the algorithm is validated for synthetic loads with known analytical solutions.We present 3D-MSM measurements of monolayer stress in micropatterned islands of endothelial cells of different sizes and shapes. These data indicate that intracellular stresses caused by lateral deformation emerge collectively over long distances; they increase with the distance from the island edge until they reach a constant value that is independent of island size. On the other hand, bending-induced intracellular stresses are more concentrated spatially and remain confined to within 1-2 cell lengths of bending sites. The magnitude of these bending stresses is highest at the edges of the cell islands, where they can exceed the intracellular stresses caused by lateral deformations. Our data from non-patterned monolayers suggests that biomechanical perturbations far away from monolayer edges also cause significant localized alterations in bending tension. The localized effect of bending-induced stresses may be important in processes like cellular extravasation, which are accompanied by significant normal deflections of a cell monolayer (i.e. the endothelium), and which require localized changes in monolayer permeability.

High precision measurement of electrical resistance across endothelial cell monolayers

1995 ◽  
Vol 430 (1) ◽  
pp. 145-147 ◽  
Author(s):  
Walter Tschugguel ◽  
Zydi Zhegu ◽  
Leszek Gajdzik ◽  
Manfred Maier ◽  
Bernd R. Binder ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
High Precision ◽  
Precision Measurement ◽  
Electrical Resistance ◽  
High Precision Measurement ◽  
Cell Monolayers

Histamine decreases the permeability of an endothelial cell monolayer by stimulating cyclic AMP production through the H2-receptor

1992 ◽  
Vol 101 (4) ◽  
pp. 745-750
Author(s):  
T. Takeda ◽  
Y. Yamashita ◽  
S. Shimazaki ◽  
Y. Mitsui
Keyword(s):  
Endothelial Cell ◽  
Cyclic Amp ◽  
Second Messengers ◽  
Cell Monolayer ◽  
Free Calcium ◽  
Endothelial Cell Monolayer ◽  
H2 Receptor ◽  
Cell Monolayers ◽  
H2 Blocker

To determine if histamine acts directly on the vascular endothelium, the effect of histamine on the permeability of cultured human endothelial cell monolayers and the role of second messengers were examined. The addition of 10(−6) to 10(−4) M histamine to the culture medium decreased the endothelial cell monolayer permeability and increased both cyclic AMP and free-calcium levels. The decrease in permeability and the increase in cyclic AMP mediated by histamine were prevented by an H2-blocker (famotidine) while the increase in free-calcium was inhibited by an H1-blocker (diphenhydramine). These results suggest that histamine decreases the permeability of endothelial cell monolayers through the H2-receptor, and cyclic AMP plays a more important role than calcium ion as a second messenger.

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