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Cited by 47Barrier Structure and Function StudiesDrug Transport and Resistance MechanismsIon Transport and Channel Regulation

Comparative studies between the murine immortalized brain endothelial cell line (bEnd.3) and induced pluripotent stem cell-derived human brain endothelial cells for paracellular transport

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Sun, Jiahong, Ou, Weijun, Han, Derick, Paganini-Hill, Annlia, Fisher, Mark J., Sumbria, Rachita K.·2022·PLOS ONE
DOI 10.1371/journal.pone.0268860PMC PMC9132315
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Abstract

Brain microvascular endothelial cells, forming the anatomical site of the blood-brain barrier (BBB), are widely used as in vitro complements to in vivo BBB studies. Among the immortalized cells used as in vitro BBB models, the murine-derived bEnd.3 cells offer culturing consistency and low cost and are well characterized for functional and transport assays, but result in low transendothelial electrical resistance (TEER). Human-induced pluripotent stem cells differentiated into brain microvascular endothelial cells (ihBMECs) have superior barrier properties, but the process of differentiation is time-consuming and can result in mixed endothelial-epithelial gene expression. Here we performed a side-by-side comparison of the ihBMECs and bEnd.3 cells for key paracellular diffusional transport characteristics. The TEER across the ihBMECs was 45- to 68-fold higher than the bEnd.3 monolayer. The ihBMECs had significantly lower tracer permeability than the bEnd.3 cells. Both, however, could discriminate between the paracellular permeabilities of two tracers: sodium fluorescein (MW: 376 Da) and fluorescein isothiocyanate (FITC)–dextran (MW: 70 kDa). FITC-dextran permeability was a strong inverse-correlate of TEER in the bEnd.3 cells, whereas sodium fluorescein permeability was a strong inverse-correlate of TEER in the ihBMECs. Both bEnd.3 cells and ihBMECs showed the typical cobblestone morphology with robust uptake of acetylated LDL and strong immuno-positivity for vWF. Both models showed strong claudin-5 expression, albeit with differences in expression location. We further confirmed the vascular endothelial- (CD31 and tube-like formation) and erythrophagocytic-phenotypes and the response to inflammatory stimuli of ihBMECs. Overall, both bEnd.3 cells and ihBMECs express key brain endothelial phenotypic markers, and despite differential TEER measurements, these in vitro models can discriminate between the passage of different molecular weight tracers. Our results highlight the need to corroborate TEER measurements with different molecular weight tracers and that the bEnd.3 cells may be suitable for large molecule transport studies despite their low TEER.

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  • These FITC-dextran transport values are consistent with other values reported in literature for in vitro BBB models and demonstrate that the monolayer possesses high structural integrity and selective permeability.

    “These values are consistent with other values reported in literature for in vitro BBB models and demonstrate that the monolayer possesses high structural integrity and selective permeability.37,38”

    From Predictive High-Throughput Platform for Dual Screening of mRNA Lipid Nanoparticle Blood–Brain Barrier Transfection and Crossing (2024) · cites this paper as [38]

    The Discussion (page not provided) says its FITC-dextran permeability values are in range with previous work, and the Results report that both monolayers restricted FITC-dextran passage. But the paper distinguishes the tighter ihBMEC barrier from the leakier bEnd.3 barrier; it does not establish that the citing paper’s unspecified monolayer has high structural integrity and selective permeability. (LLM, conf=0.85)

    Experimental — accuracy not yet measured

  • TEER was around 20 Ω/cm2 on Day 6 after cell seeding, consistent with other reported values.

    “The TEER was around 20 Ω/cm2 on Day 6 after cell seeding, consistent with other reported values (Figure S3).37,38”

    From Predictive High-Throughput Platform for Dual Screening of mRNA Lipid Nanoparticle Blood–Brain Barrier Transfection and Crossing (2024) · cites this paper as [38]

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Cited by

Sorted by citations

Authors
Sun, J., Ou, W., Han, D., Paganini-Hill, A., Fisher, M. J., & Sumbria, R. K.
Affiliations
  • Chapman University (US)
  • Keck Graduate Institute (US)
  • University of California, Irvine (US)
Corresponding author
Sumbria, Rachita K.
ORCID
Venue
PLOS ONE
Publisher
Public Library of Science (PLoS)
Published
2022-05-25
Volume
17
Issue
5
Pages
e0268860
ISSN
1932-6203 (ISSN-L)
DOI
10.1371/journal.pone.0268860
PMC
PMC9132315
OpenAlex
W4281555338
Cited by
47
References
2
Citations by year
2026: 12 · 2025: 17 · 2024: 10 · 2023: 7 · 2022: 1
FWCI
3.07
Type
journal article
Language
en
Open access
Licence
Locations
  • PLoS ONE (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • Chapman University Digital Commons (Chapman University) (submittedVersion, cc-by)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion)
  • eScholarship (California Digital Library) (submittedVersion, cc-by)
Funders
  • National Institute of Neurological Disorders and Stroke (R01NS20989)
  • National Institute on Aging (R01AG072896)
Topics
  • Life Sciences › Neuroscience › Neurology › Barrier Structure and Function Studies
  • Health Sciences › Medicine › Oncology › Drug Transport and Resistance Mechanisms
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Ion Transport and Channel Regulation
Keywords
blood-brain barrier, brain microvascular endothelial cells, bEnd.3 cells, induced pluripotent stem cell-derived endothelial cells, paracellular permeability, transendothelial electrical resistance, FITC-dextran, fluorescein, claudin-5, acetylated LDL uptake, von Willebrand factor, endothelial phenotype, inflammation, in vitro BBB model
MeSH terms
Animals, Blood-Brain Barrier, Brain, Cell Line, Cells, Cultured, Humans, Fluorescein, Endothelial Cells, Mice, Induced Pluripotent Stem Cells

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