Much of what is currently known about the role of the blood–brain barrier (BBB) in regulating the passage of chemicals from the blood stream to the central nervous system (CNS) comes from animal in vivo models (requiring extrapolation to human relevance) and 2D static in vitro systems, which fail to capture the rich cell–cell and cell–matrix interactions of the dynamic 3D in vivo tissue microenvironment. In this work we have developed a BBB platform that allows for a high degree of customization in cellular composition, cellular orientation, and physiologically-relevant fluid dynamics. The system characterized and presented in this study reproduces key characteristics of a BBB model (e.g. tight junctions, efflux pumps) allowing for the formation of a selective and functional barrier. We demonstrate that our in vitro BBB is responsive to both biochemical and mechanical cues. This model further allows for culture of a CNS-like space around the BBB. The design of this platform is a valuable tool for studying BBB function as well as for screening of novel therapeutics.
This is a preview of subscription content, access via your institution.
Buy single article
Instant access to the full article PDF.
Tax calculation will be finalised during checkout.
The data that support the findings of this study are available from the corresponding authors upon reasonable request.
Abbott, N. J., L. Rönnbäck, and E. Hansson. Astrocyte–endothelial interactions at the blood–brain barrier. Nat. Rev. Neurosci. 7:41, 2006.
Adriani, G., D. Ma, A. Pavesi, R. D. Kamm, and E. L. K. Goh. A 3D neurovascular microfluidic model consisting of neurons, astrocytes and cerebral endothelial cells as a blood–brain barrier. Lab Chip 17:448–459, 2017.
Appelt-Menzel, A., A. Cubukova, K. Günther, F. Edenhofer, J. Piontek, G. Krause, T. Stüber, H. Walles, W. Neuhaus, and M. Metzger. Establishment of a human blood–brain barrier co-culture model mimicking the neurovascular unit using induced pluri- and multipotent stem cells. Stem Cell Rep. 8:894–906, 2017.
Au-Yeung, K. L., K. Y. Sze, M. H. Sham, and B. P. Chan. Development of a micromanipulator-based loading device for mechanoregulation study of human mesenchymal stem cells in three-dimensional collagen constructs. Tissue Eng. C 16:93–107, 2010.
Bajaj, P., B. Reddy, Jr, L. Millet, C. Wei, P. Zorlutuna, G. Bao, and R. Bashir. Patterning the differentiation of C2C12 skeletal myoblasts. Integr. Biol. 3:897–909, 2011.
Bang, S., S.-R. Lee, J. Ko, K. Son, D. Tahk, J. Ahn, C. Im, and N. L. Jeon. A low permeability microfluidic blood–brain barrier platform with direct contact between perfusable vascular network and astrocytes. Sci. Rep. 7:8083, 2017.
Banks, W. A. Developing drugs that can cross the blood–brain barrier: applications to Alzheimer’s disease. BMC Neurosci. 9:S2–S2, 2008.
Biemans, E. A. L. M., L. Jäkel, R. M. W. de Waal, H. B. Kuiperij, and M. M. Verbeek. Limitations of the hCMEC/D3 cell line as a model for Aβ clearance by the human blood–brain barrier. J. Neurosci. Res. 95:1513–1522, 2017.
Birgersdotter, A., R. Sandberg, and I. Ernberg. Gene expression perturbation in vitro—a growing case for three-dimensional (3D) culture systems. Semin. Cancer Biol. 15:405–412, 2005.
Booth, R., and H. Kim. Characterization of a microfluidic in vitro model of the blood–brain barrier (muBBB). Lab Chip 12:1784–1792, 2012.
Brown, J. A., V. Pensabene, D. A. Markov, V. Allwardt, M. D. Neely, M. Shi, C. M. Britt, O. S. Hoilett, Q. Yang, B. M. Brewer, P. C. Samson, L. J. McCawley, J. M. May, D. J. Webb, D. Li, A. B. Bowman, R. S. Reiserer, and J. P. Wikswo. Recreating blood–brain barrier physiology and structure on chip: a novel neurovascular microfluidic bioreactor. Biomicrofluidics 9:054124, 2015.
Chistiakov, D. A., A. N. Orekhov, and Y. V. Bobryshev. Effects of shear stress on endothelial cells: go with the flow. Acta Physiol. 219:382–408, 2017.
Cho, C.-F., J. M. Wolfe, C. M. Fadzen, D. Calligaris, K. Hornburg, E. A. Chiocca, N. Y. R. Agar, B. L. Pentelute, and S. E. Lawler. Blood–brain-barrier spheroids as an in vitro screening platform for brain-penetrating agents. Nat. Commun. 8:15623, 2017.
Cucullo, L., N. Marchi, M. Hossain, and D. Janigro. A dynamic in vitro BBB model for the study of immune cell trafficking into the central nervous system. J. Cereb. Blood Flow Metab. 31:767–777, 2011.
Deosarkar, S. P., B. Prabhakarpandian, B. Wang, J. B. Sheffield, B. Krynska, and M. F. Kiani. A novel dynamic neonatal blood–brain barrier on a chip. PLoS ONE 10:e0142725, 2015.
DeStefano, J. G., J. J. Jamieson, R. M. Linville, and P. C. Searson. Benchmarking in vitro tissue-engineered blood–brain barrier models. Fluids Barriers CNS 15:32, 2018.
DeStefano, J. G., Z. S. Xu, A. J. Williams, N. Yimam, and P. C. Searson. Effect of shear stress on iPSC-derived human brain microvascular endothelial cells (dhBMECs). Fluids Barriers CNS 14:20, 2017.
Falanga, A. P., G. Pitingolo, M. Celentano, A. Cosentino, P. Melone, R. Vecchione, D. Guarnieri, and P. A. Netti. Shuttle-mediated nanoparticle transport across an in vitro brain endothelium under flow conditions. Biotechnol. Bioeng. 114:1087–1095, 2017.
Förster, C., M. Burek, I. A. Romero, B. Weksler, P.-O. Couraud, and D. Drenckhahn. Differential effects of hydrocortisone and TNFα on tight junction proteins in an in vitro model of the human blood–brain barrier. J. Physiol. 586:1937–1949, 2008.
Garcia, P. A., J. H. Rossmeisl, J. L. Robertson, J. D. Olson, A. J. Johnson, T. L. Ellis, and R. V. Davalos. 70-T magnetic resonance imaging characterization of acute blood–brain-barrier disruption achieved with intracranial irreversible electroporation. PLoS ONE 7:e50482, 2012.
Griep, L. M., F. Wolbers, B. de Wagenaar, P. M. ter Braak, B. B. Weksler, I. A. Romero, P. O. Couraud, I. Vermes, A. D. van der Meer, and A. van den Berg. BBB ON CHIP: microfluidic platform to mechanically and biochemically modulate blood–brain barrier function. Biomed. Microdevices 15:145–150, 2013.
Hatherell, K., P.-O. Couraud, I. A. Romero, B. Weksler, and G. J. Pilkington. Development of a three-dimensional, all-human in vitro model of the blood–brain barrier using mono-, co-, and tri-cultivation Transwell models. J. Neurosci. Methods 199:223–229, 2011.
Helms, H. C., N. J. Abbott, M. Burek, R. Cecchelli, P.-O. Couraud, M. A. Deli, C. Förster, H. J. Galla, I. A. Romero, E. V. Shusta, M. J. Stebbins, E. Vandenhaute, B. Weksler, and B. Brodin. In vitro models of the blood–brain barrier: an overview of commonly used brain endothelial cell culture models and guidelines for their use. J. Cereb. Blood Flow Metab. 36:862–890, 2016.
Herland, A., A. D. van der Meer, E. A. FitzGerald, T.-E. Park, J. J. F. Sleeboom, and D. E. Ingber. Distinct contributions of astrocytes and pericytes to neuroinflammation identified in a 3D human blood–brain barrier on a chip. PLoS ONE 11:e0150360, 2016.
Hoosain, F. G., Y. E. Choonara, L. K. Tomar, P. Kumar, C. Tyagi, L. C. du Toit, and V. Pillay. Bypassing P-glycoprotein drug efflux mechanisms: possible applications in pharmacoresistant schizophrenia therapy. BioMed Res. Int. 2015:484963, 2015.
Hynynen, K., N. McDannold, N. A. Sheikov, F. A. Jolesz, and N. Vykhodtseva. Local and reversible blood–brain barrier disruption by noninvasive focused ultrasound at frequencies suitable for trans-skull sonications. NeuroImage 24:12–20, 2005.
Jamieson, J. J., P. C. Searson, and S. Gerecht. Engineering the human blood–brain barrier in vitro. J. Biol. Eng. 11:37, 2017.
Jeong, S., S. Kim, J. Buonocore, J. Park, C. J. Welsh, J. Li, and A. Han. A three-dimensional arrayed microfluidic blood–brain barrier model with integrated electrical sensor array. IEEE Trans. Biomed. Eng. 65:431–439, 2018.
Jiang, L., S. Li, J. Zheng, Y. Li, and H. Huang. Recent progress in microfluidic models of the blood–brain barrier. Micromachines (Basel) 2019. https://doi.org/10.3390/mi10060375.
Kalvass, J. C., J. W. Polli, D. L. Bourdet, B. Feng, S.-M. Huang, X. Liu, Q. R. Smith, L. K. Zhang, and M. J. Zamek-Gliszczynski. Why clinical modulation of efflux transport at the human blood–brain barrier is unlikely: the ITC evidence-based position. Clin. Pharmacol. Ther. 94:80–94, 2013.
Koo, Y., B. T. Hawkins, and Y. Yun. Three-dimensional (3D) tetra-culture brain on chip platform for organophosphate toxicity screening. Sci. Rep. 8:2841, 2018.
Liu, C., X.-N. Liu, G.-L. Wang, Y. Hei, S. Meng, L.-F. Yang, L. Yuan, and Y. Xie. A dual-mediated liposomal drug delivery system targeting the brain: rational construction, integrity evaluation across the blood–brain barrier, and the transporting mechanism to glioma cells. Int. J. Nanomed. 12:2407–2425, 2017.
Mairey, E., A. Genovesio, E. Donnadieu, C. Bernard, F. Jaubert, E. Pinard, J. Seylaz, J. C. Olivo-Marin, X. Nassif, and G. Dumenil. Cerebral microcirculation shear stress levels determine Neisseria meningitidis attachment sites along the blood–brain barrier. J. Exp. Med. 203:1939–1950, 2006.
Man, S., E. E. Ubogu, K. A. Williams, B. Tucky, M. K. Callahan, and R. M. Ransohoff. Human brain microvascular endothelial cells and umbilical vein endothelial cells differentially facilitate leukocyte recruitment and utilize chemokines for T cell migration. Clin. Dev. Immunol. 2008:384982, 2008.
Minagar, A., and J. S. Alexander. Blood–brain barrier disruption in multiple sclerosis. Mult. Scler. J. 9:540–549, 2003.
Nakagawa, S., M. A. Deli, H. Kawaguchi, T. Shimizudani, T. Shimono, Á. Kittel, K. Tanaka, and M. Niwa. A new blood–brain barrier model using primary rat brain endothelial cells, pericytes and astrocytes. Neurochem. Int. 54:253–263, 2009.
Nguyen, T. P. T., B. M. Tran, and N. Y. Lee. Microfluidic approach for the fabrication of cell-laden hollow fibers for endothelial barrier research. J. Mater. Chem. B 6:6057–6066, 2018.
Ni, Y., T. Teng, R. Li, A. Simonyi, G. Y. Sun, and J. C. Lee. TNFα alters occludin and cerebral endothelial permeability: role of p38MAPK. PLoS ONE 12:e0170346, 2017.
Odijk, M., A. D. Van Der Meer, D. Levner, H. J. Kim, M. W. Van Der Helm, L. I. Segerink, J. P. Frimat, G. A. Hamilton, D. E. Ingber, and A. Van Den Berg. Measuring direct current trans-epithelial electrical resistance in organ-on-a-chip microsystems. Lab Chip 15:745–752, 2015.
Pardridge, W. M. The blood–brain barrier: bottleneck in brain drug development. NeuroRx 2:3–14, 2005.
Partyka, P. P., G. A. Godsey, J. R. Galie, M. C. Kosciuk, N. K. Acharya, R. G. Nagele, and P. A. Galie. Mechanical stress regulates transport in a compliant 3D model of the blood–brain barrier. Biomaterials 115:30–39, 2017.
Persidsky, Y., S. H. Ramirez, J. Haorah, and G. D. Kanmogne. Blood–brain barrier: structural components and function under physiologic and pathologic conditions. J. Neuroimmune Pharmacol. 1:223–236, 2006.
Prabhakarpandian, B., M.-C. Shen, J. B. Nichols, I. R. Mills, M. Sidoryk-Wegrzynowicz, M. Aschner, and K. Pant. SyM-BBB: a microfluidic blood brain barrier model. Lab Chip 13:1093–1101, 2013.
Reinitz, A., J. DeStefano, M. Ye, A. D. Wong, and P. C. Searson. Human brain microvascular endothelial cells resist elongation due to shear stress. Microvasc. Res. 99:8–18, 2015.
Saunders, N. R., M. D. Habgood, K. Møllgård, and K. M. Dziegielewska. The biological significance of brain barrier mechanisms: help or hindrance in drug delivery to the central nervous system? F1000Research 2016. https://doi.org/10.12688/f1000research.7378.1.
Sellgren, K. L., B. T. Hawkins, and S. Grego. An optically transparent membrane supports shear stress studies in a three-dimensional microfluidic neurovascular unit model. Biomicrofluidics 9:061102, 2015.
Shimokawa, H., and S. Godo. Diverse functions of endothelial NO synthases system: NO and EDH. J. Cardiovasc. Pharmacol. 67:361–366, 2016.
Stanness, K. A., L. E. Westrum, E. Fornaciari, P. Mascagni, J. A. Nelson, S. G. Stenglein, T. Myers, and D. Janigro. Morphological and functional characterization of an in vitro blood–brain barrier model. Brain Res. 771:329–342, 1997.
Sweeney, M. D., A. P. Sagare, and B. V. Zlokovic. Blood–brain barrier breakdown in Alzheimer disease and other neurodegenerative disorders. Nat. Rev. Neurol. 14:133, 2018.
Tibbe, M. P., A. M. Leferink, A. van den Berg, J. C. T. Eijkel, and L. I. Segerink. Microfluidic gel patterning method by use of a temporary membrane for organ-on-chip applications. Adv. Mater. Technol. 3:1700200, 2018.
Urich, E., C. Patsch, S. Aigner, M. Graf, R. Iacone, and P.-O. Freskgård. Multicellular self-assembled spheroidal model of the blood brain barrier. Sci. Rep. 3:1500, 2013.
van der Helm, M. W., A. D. van der Meer, J. C. T. Eijkel, A. van den Berg, and L. I. Segerink. Microfluidic organ-on-chip technology for blood–brain barrier research. Tissue Barriers 4:e1142493, 2016.
Walsby, E., A. Buggins, S. Devereux, C. Jones, G. Pratt, P. Brennan, C. Fegan, and C. Pepper. Development and characterization of a physiologically relevant model of lymphocyte migration in chronic lymphocytic leukemia. Blood 123:3607–3617, 2014.
Wang, Y. I., H. E. Abaci, and M. L. Shuler. Microfluidic blood–brain barrier model provides in vivo-like barrier properties for drug permeability screening. Biotechnol. Bioeng. 114:184–194, 2017.
Wang, J. D., E.-S. Khafagy, K. Khanafer, S. Takayama, and M. E. H. ElSayed. Organization of endothelial cells, pericytes, and astrocytes into a 3D microfluidic in vitro model of the blood–brain barrier. Mol. Pharm. 13:895–906, 2016.
Wei, X., X. Chen, M. Ying, and W. Lu. Brain tumor-targeted drug delivery strategies. Acta Pharm. Sin. B 4:193–201, 2014.
Weksler, B., I. A. Romero, and P.-O. Couraud. The hCMEC/D3 cell line as a model of the human blood brain barrier. Fluids Barriers CNS 10:16–16, 2013.
Ye, M., H. M. Sanchez, M. Hultz, Z. Yang, M. Bogorad, A. D. Wong, and P. C. Searson. Brain microvascular endothelial cells resist elongation due to curvature and shear stress. Sci. Rep. 4:4681, 2014.
Zhao, C., H. Wang, C. Xiong, and Y. Liu. Hypoxic glioblastoma release exosomal VEGF-A induce the permeability of blood–brain barrier. Biochem. Biophys. Res. Commun. 502:324–331, 2018.
Zlokovic, B. V. The blood–brain barrier in health and chronic neurodegenerative disorders. Neuron 57:178–201, 2008.
This work was funded by LDRD Awards 14-SI-001 and 17-SI-002 under the auspices of the U.S. Department of Energy by Lawrence Livermore National Laboratory under Contract DE-AC52-07NA27344 Lawrence Livermore National Security, LLC. We acknowledge Scott Erickson, Sierra Levenson, Jonathan Adorno, and Haley Sandvik for their help with device fabrication efforts. LLNL-JRNL-758697.
Conflict of interests
The authors declare no competing interests.
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Associate Editor Debra T. Auguste oversaw the review of this article.
About this article
Cite this article
Moya, M.L., Triplett, M., Simon, M. et al. A Reconfigurable In Vitro Model for Studying the Blood–Brain Barrier. Ann Biomed Eng 48, 780–793 (2020). https://doi.org/10.1007/s10439-019-02405-y
- Blood–brain barrier
- Endothelial cells