scholarly journals In Vitro Methods for Measuring the Permeability of Cell Monolayers

2022 ◽  
Author(s):  
Radoslaw Bednarek
Keyword(s):  
Impedance Measurement ◽  
Endothelial Permeability ◽  
Intercellular Junctions ◽  
In Vitro Models ◽  
Leukocyte Transmigration ◽  
Cell Monolayers ◽  
Electrical Impedance Measurement ◽  
In Vitro Systems ◽  
Large Groups

Cell monolayers, including endothelial and epithelial cells, play crucial roles in regulating the transport of biomolecules to underlying tissues and structures via intercellular junctions. Moreover, the monolayers form a semipermeable barrier across which leukocyte transmigration is tightly regulated. The inflammatory cytokines can disrupt the epithelial and endothelial permeability, thus the reduced barrier integrity is a hallmark of epithelial and endothelial dysfunction related with numerous pathological conditions, including cancer-related inflammation. Therefore, the assessment of barrier function is critical in in vitro models of barrier-forming tissues. This review summarizes the commercially available in vitro systems used to measure the permeability of cellular monolayers. The presented techniques are separated in two large groups: macromolecular tracer flux assays, and electrical impedance measurement-based permeability assays. The presented techniques are briefly described and compared.

scholarly journals Human Oligodendrocytes and Myelin In Vitro to Evaluate Developmental Neurotoxicity

2021 ◽  
Vol 22 (15) ◽  
pp. 7929
Author(s):  
Megan Chesnut ◽  
Thomas Hartung ◽  
Helena Hogberg ◽  
David Pamies
Keyword(s):  
Large Scale ◽  
In Vitro Models ◽  
Environmental Chemicals ◽  
Developmental Neurotoxicity ◽  
In Vivo Studies ◽  
Regulatory Guidelines ◽  
In Vitro Systems ◽  
Human Oligodendrocytes

Neurodevelopment is uniquely sensitive to toxic insults and there are concerns that environmental chemicals are contributing to widespread subclinical developmental neurotoxicity (DNT). Increased DNT evaluation is needed due to the lack of such information for most chemicals in common use, but in vivo studies recommended in regulatory guidelines are not practical for the large-scale screening of potential DNT chemicals. It is widely acknowledged that developmental neurotoxicity is a consequence of disruptions to basic processes in neurodevelopment and that testing strategies using human cell-based in vitro systems that mimic these processes could aid in prioritizing chemicals with DNT potential. Myelination is a fundamental process in neurodevelopment that should be included in a DNT testing strategy, but there are very few in vitro models of myelination. Thus, there is a need to establish an in vitro myelination assay for DNT. Here, we summarize the routes of myelin toxicity and the known models to study this particular endpoint.

Protein, not adenosine or adenine nucleotides, mediates platelet decrease in endothelial permeability

1997 ◽  
Vol 273 (5) ◽  
pp. H2304-H2311 ◽  
Author(s):  
Sandeep Patil ◽  
John E. Kaplan ◽  
Fred L. Minnear
Keyword(s):  
High Performance ◽  
Adenine Nucleotides ◽  
Endothelial Permeability ◽  
Platelet Response ◽  
Cell Monolayers ◽  
Adenosine Receptor Antagonist ◽  
Albumin Clearance ◽  
Protein Permeability

Platelets and platelet-conditioned medium (PCM) decrease endothelial protein permeability in vitro. Adenosine and a >100-kDa protein have previously been implicated as the soluble factors released from platelets that decrease endothelial permeability. The objective of this study was to further investigate the role of adenosine in this platelet response. Measurements of adenosine and its precursor adenine nucleotides by high-performance liquid chromatography were correlated with the assessment of permeability by125I-labeled albumin clearance and electrical resistance across endothelial cell monolayers derived from the bovine pulmonary artery. PCM contained micromolar concentrations of AMP, ADP, and ATP, but adenosine was below detectable levels (≤0.1 μM). Adenosine deaminase, an enzyme that converts adenosine to inactive inosine, or an adenosine-receptor antagonist did not block the platelet- or PCM-mediated decrease in endothelial permeability. A <3-kDa fraction of PCM that contained micromolar concentrations of AMP and ADP did not affect endothelial permeability, whereas a >3-kDa fraction that contained much reduced levels of AMP and ADP significantly decreased permeability. This activity of PCM was sensitive to insoluble trypsin. This study rules out adenosine and adenine nucleotides as primary factors in the platelet-induced decrease in endothelial permeability and suggests that the active factor is a protein.

Reciprocal regulation of endothelin-1 and endothelial constitutive NOS in proliferating endothelial cells

1995 ◽  
Vol 269 (6) ◽  
pp. H1988-H1997 ◽  
Author(s):  
M. A. Flowers ◽  
Y. Wang ◽  
R. J. Stewart ◽  
B. Patel ◽  
P. A. Marsden
Keyword(s):  
Endothelial Cell ◽  
In Vitro Models ◽  
Endothelial Cell Proliferation ◽  
Mrna Levels ◽  
Proliferating Cells ◽  
Reciprocal Regulation ◽  
Cell Monolayers ◽  
Endothelin 1 ◽  
Mrna Transcripts

The expression of endothelin-1 (ET-1) and endothelial constitutive nitric oxide synthase (ecNOS) was assessed in two independent in vitro models: asynchronously differentially proliferating cultures and wounded endothelial cell monolayers. Northern blot analysis of RNA isolated from preconfluent, confluent, and postconfluent cells revealed a fourfold rise in ET-1 mRNA transcripts, whereas levels of ecNOS mRNA transcripts were reduced twofold in proliferating cells. Nuclear run-off analysis demonstrated that increased steady-state ET-1 mRNA content in proliferating cells was mediated, in part, by increased gene transcription. In contrast, ecNOS transcription rates in proliferating cells were not decreased compared with quiescent nonproliferating cells, indicating that mRNA destabilization mediated the decreased ecNOS mRNA levels in proliferating endothelium. Concordant changes in protein expression were documented for both ET-1 and ecNOS. In injured endothelial cell monolayers, in situ cRNA hybridization demonstrated increased mRNA transcript levels for ET-1 in growth fronts of injured endothelial monolayers. These data are taken to indicate that expression of ET-1 and ecNOS is reciprocally regulated in two different models of endothelial cell proliferation.

scholarly journals Regenerative Neurology and Regenerative Cardiology: Shared Hurdles and Achievements

2022 ◽  
Vol 23 (2) ◽  
pp. 855
Author(s):  
Dinko Mitrečić ◽  
Valentina Hribljan ◽  
Denis Jagečić ◽  
Jasmina Isaković ◽  
Federica Lamberto ◽  
...  
Keyword(s):  
Biomedical Research ◽  
Cell Types ◽  
In Vitro Models ◽  
Medical Systems ◽  
Heart Infarction ◽  
Affected Tissue ◽  
In Vitro Systems ◽  
Different Cell Types ◽  
The Brain

From the first success in cultivation of cells in vitro, it became clear that developing cell and/or tissue specific cultures would open a myriad of new opportunities for medical research. Expertise in various in vitro models has been developing over decades, so nowadays we benefit from highly specific in vitro systems imitating every organ of the human body. Moreover, obtaining sufficient number of standardized cells allows for cell transplantation approach with the goal of improving the regeneration of injured/disease affected tissue. However, different cell types bring different needs and place various types of hurdles on the path of regenerative neurology and regenerative cardiology. In this review, written by European experts gathered in Cost European action dedicated to neurology and cardiology-Bioneca, we present the experience acquired by working on two rather different organs: the brain and the heart. When taken into account that diseases of these two organs, mostly ischemic in their nature (stroke and heart infarction), bring by far the largest burden of the medical systems around Europe, it is not surprising that in vitro models of nervous and heart muscle tissue were in the focus of biomedical research in the last decades. In this review we describe and discuss hurdles which still impair further progress of regenerative neurology and cardiology and we detect those ones which are common to both fields and some, which are field-specific. With the goal to elucidate strategies which might be shared between regenerative neurology and cardiology we discuss methodological solutions which can help each of the fields to accelerate their development.

Comparative Cell Toxicology: The Basis for In Vitro Toxicity Testing

1994 ◽  
Vol 22 (3) ◽  
pp. 168-174
Author(s):  
Hasso Seibert ◽  
Michael Gulden ◽  
Jens-Uwe Voss
Keyword(s):  
Toxicity Testing ◽  
Cell Types ◽  
Scientific Basis ◽  
In Vitro Models ◽  
In Vitro Toxicity ◽  
Full Potential ◽  
Type B ◽  
Cell Type ◽  
In Vitro Systems

If “cell toxicology” is defined as the discipline aimed at studying the general principles of chemical interference with cellular structures and/or functions, then “comparative cell toxicology” may be defined as the study of the variety of responses to xenobiotics using: (a) different endpoints within one cell type; (b) cell types from different tissues from one species; and (c) homologous cell types from different species. If the full potential of in vitro models for toxicity testing is to be realised and the scientific basis for hazard assessment improved, then comparative cell toxicological approaches have to be developed further. In the present paper, an approach using different in vitro systems is described. The approach is aimed at the assessment of the basic toxicological characteristics of chemicals.

scholarly journals Evaluation of Lipid Accumulation Using Electrical Impedance Measurement under Three-Dimensional Culture Condition

Micromachines ◽  
2019 ◽  
Vol 10 (7) ◽  
pp. 455 ◽  
Author(s):  
Daiki Zemmyo ◽  
Shogo Miyata
Keyword(s):  
Lipid Accumulation ◽  
High Frequency ◽  
Culture System ◽  
Electrical Impedance ◽  
Three Dimensional ◽  
Impedance Measurement ◽  
Electrical Impedance Measurement ◽  
The Relationship ◽  
Dimensional Cell

The degeneration of adipocyte has been reported to cause obesity, metabolic syndrome, and other diseases. To treat these diseases, an effective in vitro evaluation and drug-screening system for adipocyte culture is required. The objective of this study is to establish an in vitro three-dimensional cell culture system to enable the monitoring of lipid accumulation by measuring electrical impedance, and to determine the relationship between the impedance and lipid accumulation of adipocytes cultured three dimensionally. Consequently, pre-adipocytes, 3T3-L1 cells, were cultured and differentiated to the adipocytes in our culture system, and the electrical impedance of the three-dimensional adipocyte culture at a high frequency was related to the lipid accumulation of the adipocytes. In conclusion, the lipid accumulation of adipocytes could be evaluated in real time by monitoring the electrical impedance during in vitro culture.

scholarly journals Senescence and associated blood–brain barrier alterations in vitro

2021 ◽  
Author(s):  
Ellaine Salvador ◽  
Malgorzata Burek ◽  
Mario Löhr ◽  
Michiaki Nagai ◽  
Carsten Hagemann ◽  
...  
Keyword(s):  
Blood Brain Barrier ◽  
Intercellular Junctions ◽  
In Vitro Models ◽  
Brain Barrier ◽  
Initial Attempt ◽  
Blood Brain ◽  
The Central Nervous System ◽  
Vitro Model ◽  
The Relationship

AbstractProgressive deterioration of the central nervous system (CNS) is commonly associated with aging. An important component of the neurovasculature is the blood–brain barrier (BBB), majorly made up of endothelial cells joined together by intercellular junctions. The relationship between senescence and changes in the BBB has not yet been thoroughly explored. Moreover, the lack of in vitro models for the study of the mechanisms involved in those changes impede further and more in-depth investigations in the field. For this reason, we herein present an in vitro model of the senescent BBB and an initial attempt to identify senescence-associated alterations within.

scholarly journals Review: Challenges of In Vitro CAF Modelling in Liver Cancers

Cancers ◽  
2021 ◽  
Vol 13 (23) ◽  
pp. 5914
Author(s):  
Alba Herrero ◽  
Elisabeth Knetemann ◽  
Inge Mannaerts
Keyword(s):  
Liver Cancer ◽  
Treatment Strategies ◽  
3D Models ◽  
Tumour Microenvironment ◽  
Current Treatment ◽  
In Vitro Models ◽  
Liver Cancers ◽  
In Vitro Systems ◽  
The Impact

Primary and secondary liver cancer are the third cause of death in the world, and as the incidence is increasing, liver cancer represents a global health burden. Current treatment strategies are insufficient to permanently cure patients from this devastating disease, and therefore other approaches are under investigation. The importance of cancer-associated fibroblasts (CAFs) in the tumour microenvironment is evident, and many pre-clinical studies have shown increased tumour aggressiveness in the presence of CAFs. However, it remains unclear how hepatic stellate cells are triggered by the tumour to become CAFs and how the recently described CAF subtypes originate and orchestrate pro-tumoural effects. Specialized in vitro systems will be needed to address these questions. In this review, we present the currently used in vitro models to study CAFs in primary and secondary liver cancer and highlight the trend from using oversimplified 2D culture systems to more complex 3D models. Relatively few studies report on the impact of cancer (sub)types on CAFs and the tumour microenvironment, and most studies investigated the impact of secreted factors due to the nature of the models.

scholarly journals Key Challenges and Opportunities Associated with the Use of In Vitro Models to Detect Human DILI: Integrated Risk Assessment and Mitigation Plans

2016 ◽  
Vol 2016 ◽  
pp. 1-20 ◽  
Author(s):  
Franck A. Atienzar ◽  
Eric A. Blomme ◽  
Minjun Chen ◽  
Philip Hewitt ◽  
J. Gerry Kenna ◽  
...  
Keyword(s):  
Risk Assessment ◽  
Data Interpretation ◽  
In Vitro Models ◽  
Drug Induced ◽  
Integrated Risk Assessment ◽  
Integrated Risk ◽  
Challenges And Opportunities ◽  
Black Box Warnings ◽  
In Vitro Systems

Drug-induced liver injury (DILI) is a major cause of late-stage clinical drug attrition, market withdrawal, black-box warnings, and acute liver failure. Consequently, it has been an area of focus for toxicologists and clinicians for several decades. In spite of considerable efforts, limited improvements in DILI prediction have been made and efforts to improve existing preclinical models or develop new test systems remain a high priority. While prediction of intrinsic DILI has improved, identifying compounds with a risk for idiosyncratic DILI (iDILI) remains extremely challenging because of the lack of a clear mechanistic understanding and the multifactorial pathogenesis of idiosyncratic drug reactions. Well-defined clinical diagnostic criteria and risk factors are also missing. This paper summarizes key data interpretation challenges, practical considerations, model limitations, and the need for an integrated risk assessment. As demonstrated through selected initiatives to address other types of toxicities, opportunities exist however for improvement, especially through better concerted efforts at harmonization of current, emerging and novel in vitro systems or through the establishment of strategies for implementation of preclinical DILI models across the pharmaceutical industry. Perspectives on the incorporation of newer technologies and the value of precompetitive consortia to identify useful practices are also discussed.

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