Blood–Brain Barrier Dynamic Device with Uniform Shear Stress Distribution for Microscopy and Permeability Measurements
Abstract
:1. Introduction
- A robust, low-cost, disposable (to limit potential contaminations or substance releases), and non-removable (to ensure sealing) barrier device,
- Cultivation of a monolayer of human BMEC on a porous membrane at the interface between two compartments,
- Simple cell culture in both static and dynamic conditions with uniform shear stress to achieve BMEC differentiation,
- Compatibility with phase contrast microscopy to visualize the entire cell monolayer for monitoring cell growth and confluence, as well as confocal microscopy for characterization,
- The possibility to combine several conditions in parallel, and to sample both compartments by recovering sufficient volumes for permeability or transporter studies.
2. Materials and Methods
2.1. Materials
2.2. Barrier Device Fabrication
2.3. Endothelial Cell Culture
2.4. Cell Culture Characterization
- Viability test:
- Immunostaining:
2.5. Permeability Measurement
: cumulative LY concentrations | : membrane surface area |
: lower compartment volume | : Initial LY concentration |
: incubation time |
3. Results
3.1. Barrier Device Design
3.1.1. Materials for Device Fabrication
3.1.2. Channel Dimensions
- Channel lengths:
- Channel width (w):
- Upper channel height (h):
- Lower channel height:
3.2. Flow Profile
- Pump system:
- Flow rate:
- Laminarity:
: culture medium density (993 kg·m−3) | : dynamic viscosity (0.72 × 10−3 kg·m−1·s−1) |
: flow velocity (in m·s−1) | : flow rate (in m3·s−1) |
: channel height (0.5 × 10−3 m) | : channel width (5 × 10−3 m) |
3.3. Cell Culture
3.4. Validation of Specific Junctional Markers
3.5. Permeability Study
4. Discussion
- Limitations and future works:
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Effective Culture Volumes and Times: | ||||
---|---|---|---|---|
Material for Culture | Cell Seeding Surface | Volume | Medium Height | Interval between Two Medium Changes |
cm² | mL | cm | Hours | |
T75 Flask | 75 | 15 * | 0.2 | 96 |
T25 Flask | 25 | 5 * | 0.2 | 96 |
Barrier device with h = 0.6 mm | 3.26 | 0.196 | 0.06 | 29 |
Barrier device with h = 0.5 mm | 3.26 | 0.163 | 0.05 | 24 |
Barrier device with h = 0.3 mm | 3.26 | 0.098 | 0.03 | 14 |
Culture Condition | Recovery (%) | Papp (10−6 cm·s−1) |
---|---|---|
Cells in static culture | 87.3 0.36 | 14.2 0.66 |
Cells in dynamic culture | 86.6 2.99 | 10.2 1.11 * |
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Choublier, N.; Müller, Y.; Gomez Baisac, L.; Laedermann, J.; de Rham, C.; Declèves, X.; Roux, A. Blood–Brain Barrier Dynamic Device with Uniform Shear Stress Distribution for Microscopy and Permeability Measurements. Appl. Sci. 2021, 11, 5584. https://doi.org/10.3390/app11125584
Choublier N, Müller Y, Gomez Baisac L, Laedermann J, de Rham C, Declèves X, Roux A. Blood–Brain Barrier Dynamic Device with Uniform Shear Stress Distribution for Microscopy and Permeability Measurements. Applied Sciences. 2021; 11(12):5584. https://doi.org/10.3390/app11125584
Chicago/Turabian StyleChoublier, Nina, Yoann Müller, Loris Gomez Baisac, Jeremy Laedermann, Casimir de Rham, Xavier Declèves, and Adrien Roux. 2021. "Blood–Brain Barrier Dynamic Device with Uniform Shear Stress Distribution for Microscopy and Permeability Measurements" Applied Sciences 11, no. 12: 5584. https://doi.org/10.3390/app11125584
APA StyleChoublier, N., Müller, Y., Gomez Baisac, L., Laedermann, J., de Rham, C., Declèves, X., & Roux, A. (2021). Blood–Brain Barrier Dynamic Device with Uniform Shear Stress Distribution for Microscopy and Permeability Measurements. Applied Sciences, 11(12), 5584. https://doi.org/10.3390/app11125584