scholarly journals A Bioinspired in vitro Lung Model to Study Particokinetics of Nano-/Microparticles Under Cyclic Stretch and Air-Liquid Interface Conditions

2021 ◽  
Vol 9 ◽  
Author(s):  
Ali Doryab ◽  
Mehmet Berat Taskin ◽  
Philipp Stahlhut ◽  
Andreas Schröppel ◽  
Sezer Orak ◽  
...  
Keyword(s):  
Elastic Modulus ◽  
Epithelial Cells ◽  
Cellular Uptake ◽  
Liquid Interface ◽  
Alveolar Epithelial Cells ◽  
Lung Model ◽  
Cyclic Stretch ◽  
Pulmonary Epithelial Cells ◽  
Air Liquid Interface

Evolution has endowed the lung with exceptional design providing a large surface area for gas exchange area (ca. 100 m2) in a relatively small tissue volume (ca. 6 L). This is possible due to a complex tissue architecture that has resulted in one of the most challenging organs to be recreated in the lab. The need for realistic and robust in vitro lung models becomes even more evident as causal therapies, especially for chronic respiratory diseases, are lacking. Here, we describe the Cyclic InVItroCell-stretch (CIVIC) “breathing” lung bioreactor for pulmonary epithelial cells at the air-liquid interface (ALI) experiencing cyclic stretch while monitoring stretch-related parameters (amplitude, frequency, and membrane elastic modulus) under real-time conditions. The previously described biomimetic copolymeric BETA membrane (5 μm thick, bioactive, porous, and elastic) was attempted to be improved for even more biomimetic permeability, elasticity (elastic modulus and stretchability), and bioactivity by changing its chemical composition. This biphasic membrane supports both the initial formation of a tight monolayer of pulmonary epithelial cells (A549 and 16HBE14o−) under submerged conditions and the subsequent cell-stretch experiments at the ALI without preconditioning of the membrane. The newly manufactured versions of the BETA membrane did not improve the characteristics of the previously determined optimum BETA membrane (9.35% PCL and 6.34% gelatin [w/v solvent]). Hence, the optimum BETA membrane was used to investigate quantitatively the role of physiologic cyclic mechanical stretch (10% linear stretch; 0.33 Hz: light exercise conditions) on size-dependent cellular uptake and transepithelial transport of nanoparticles (100 nm) and microparticles (1,000 nm) for alveolar epithelial cells (A549) under ALI conditions. Our results show that physiologic stretch enhances cellular uptake of 100 nm nanoparticles across the epithelial cell barrier, but the barrier becomes permeable for both nano- and micron-sized particles (100 and 1,000 nm). This suggests that currently used static in vitro assays may underestimate cellular uptake and transbarrier transport of nanoparticles in the lung.

scholarly journals Real-time measurement of cellular bioenergetics in fully differentiated human nasal epithelial cells grown at air-liquid-interface

2020 ◽  
Vol 318 (6) ◽  
pp. L1158-L1164
Author(s):  
Emily Mavin ◽  
Bernard Verdon ◽  
Sean Carrie ◽  
Vinciane Saint-Criq ◽  
Jason Powell ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Real Time ◽  
Aerobic Glycolysis ◽  
Airway Epithelial Cells ◽  
Respiratory Epithelium ◽  
Liquid Interface ◽  
Human Nasal Epithelial Cells ◽  
Respiratory Epithelial ◽  
Air Liquid Interface

Shifts in cellular metabolic phenotypes have the potential to cause disease-driving processes in respiratory disease. The respiratory epithelium is particularly susceptible to metabolic shifts in disease, but our understanding of these processes is limited by the incompatibility of the technology required to measure metabolism in real-time with the cell culture platforms used to generate differentiated respiratory epithelial cell types. Thus, to date, our understanding of respiratory epithelial metabolism has been restricted to that of basal epithelial cells in submerged culture, or via indirect end point metabolomics readouts in lung tissue. Here we present a novel methodology using the widely available Seahorse Analyzer platform to monitor real-time changes in the cellular metabolism of fully differentiated primary human airway epithelial cells grown at air-liquid interface (ALI). We show increased glycolytic, but not mitochondrial, ATP production rates in response to physiologically relevant increases in glucose availability. We also show that pharmacological inhibition of lactate dehydrogenase is able to reduce glucose-induced shifts toward aerobic glycolysis. This method is timely given the recent advances in our understanding of new respiratory epithelial subtypes that can only be observed in vitro through culture at ALI and will open new avenues to measure real-time metabolic changes in healthy and diseased respiratory epithelium, and in turn the potential for the development of novel therapeutics targeting metabolic-driven disease phenotypes.

scholarly journals A Custom-Made Device for Reproducibly Depositing Pre-metered Doses of Nebulized Drugs on Pulmonary Cells in vitro

2021 ◽  
Vol 9 ◽  
Author(s):  
Justus C. Horstmann ◽  
Chelsea R. Thorn ◽  
Patrick Carius ◽  
Florian Graef ◽  
Xabier Murgia ◽  
...  
Keyword(s):  
Cell Cultures ◽  
Pharmaceutical Formulations ◽  
Barrier Properties ◽  
Liquid Interface ◽  
Bronchial Epithelial ◽  
Pulmonary Epithelial Cells ◽  
Custom Made ◽  
Air Liquid Interface ◽  
Phosphate Buffered Saline

The deposition of pre-metered doses (i.e., defined before and not after exposition) at the air–liquid interface of viable pulmonary epithelial cells remains an important but challenging task for developing aerosol medicines. While some devices allow quantification of the deposited dose after or during the experiment, e.g., gravimetrically, there is still no generally accepted way to deposit small pre-metered doses of aerosolized drugs or pharmaceutical formulations, e.g., nanomedicines. Here, we describe a straightforward custom-made device, allowing connection to commercially available nebulizers with standard cell culture plates. Designed to tightly fit into the approximately 12-mm opening of either a 12-well Transwell® insert or a single 24-well plate, a defined dose of an aerosolized liquid can be directly deposited precisely and reproducibly (4.8% deviation) at the air–liquid interface (ALI) of pulmonary cell cultures. The deposited dose can be controlled by the volume of the nebulized solution, which may vary in a range from 20 to 200 μl. The entire nebulization-deposition maneuver is completed after 30 s and is spatially homogenous. After phosphate-buffered saline (PBS) deposition, the viability and barrier properties transepithelial electrical resistance (TEER) of human bronchial epithelial Calu-3 cells were not negatively affected. Straightforward in manufacture and use, the device enables reproducible deposition of metered doses of aerosolized drugs to study the interactions with pulmonary cell cultures grown at ALI conditions.

Murine Tracheal and Nasal Septal Epithelium for Air–Liquid Interface Cultures: A Comparative Study

2007 ◽  
Vol 21 (5) ◽  
pp. 533-537 ◽  
Author(s):  
Bradford A. Woodworth ◽  
Marcelo B. Antunes ◽  
Geeta Bhargave ◽  
James N. Palmer ◽  
Noam A. Cohen
Keyword(s):  
Epithelial Cells ◽  
Respiratory Epithelium ◽  
Liquid Interface ◽  
Respiratory Epithelial Cells ◽  
Cell Counts ◽  
Total Cell Population ◽  
Respiratory Epithelial ◽  
Air Liquid Interface ◽  
Ciliated Epithelial Cells

Background Air–liquid interface cultures using murine tracheal respiratory epithelium have revolutionized the in vitro study of airway diseases. However, these cultures often are impractical because of the small number of respiratory epithelial cells that can be isolated from the mouse trachea. The ability to study ciliary physiology in vitro is of utmost importance in the research of chronic rhinosinusitis (CRS). Our hypothesis is that the murine nasal septum is a better source of ciliated respiratory epithelium to develop respiratory epithelial air–liquid interface models. Methods Nasal septa and tracheas were harvested from 10 BALB/c mice. The nasal septa were harvested by using a simple and straightforward novel technique. Scanning electron microscopy was performed on all specimens. Cell counts of ciliated respiratory epithelial cells were performed at one standard magnification (1535×). Comparative analysis of proximal and distal trachea, midanterior and midposterior nasal septal epithelium, was performed. Results Independent cell counts revealed highly significant differences in the proportion of cell populations (p < 0.00001). Ciliated cell counts for the trachea (106.9 ± 28) were an average of 38.7% of the total cell population. Nasal septal ciliated epithelial cells (277.5 ± 16) comprised 90.1% of the total cell population. Conclusion To increase the yield of respiratory epithelial cells harvested from mice, we have found that the nasal septum is a far superior source when compared with the trachea. The greater surface area and increased concentration of ciliated epithelial cells has the potential to provide an eightfold increase in epithelial cells for the development of air–liquid interface cultures.

scholarly journals A Biomimetic, Copolymeric Membrane for Cell‐Stretch Experiments with Pulmonary Epithelial Cells at the Air‐Liquid Interface

2020 ◽  
pp. 2004707
Author(s):  
Ali Doryab ◽  
Mehmet Berat Taskin ◽  
Philipp Stahlhut ◽  
Andreas Schröppel ◽  
Darcy E. Wagner ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Liquid Interface ◽  
Pulmonary Epithelial Cells ◽  
Cell Stretch ◽  
Air Liquid Interface

Alveolar epithelial cells (A549) exposed at the air–liquid interface to diesel exhaust: First study in TNO’s powertrain test center

2013 ◽  
Vol 27 (8) ◽  
pp. 2342-2349 ◽  
Author(s):  
Ingeborg M. Kooter ◽  
Marcel J. Alblas ◽  
Aleksandra D. Jedynska ◽  
Maaike Steenhof ◽  
Marc M.G. Houtzager ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Diesel Exhaust ◽  
Liquid Interface ◽  
Alveolar Epithelial Cells ◽  
Alveolar Epithelial ◽  
Air Liquid Interface
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