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A face-to-face comparison of claudin-5 transduced human brain endothelial (hCMEC/D3) cells with porcine brain endothelial cells as blood-brain barrier models for drug transport studies.

Fluids Barriers CNS. 2020 Aug 26;17(1):53
Birthe Gericke 1 , Kerstin Römermann 1 , Andreas Noack 1 , Sandra Noack 2 , Jessica Kronenberg 1 , Ingolf Ernst Blasig 3 , Wolfgang Löscher 4
Birthe Gericke 1 , Kerstin Römermann 1 , Andreas Noack 1 , Sandra Noack 2 , Jessica Kronenberg 1 , Ingolf Ernst Blasig 3 , Wolfgang Löscher 4
+ et al

[No authors listed]

Author information
  • 1 Department of Pharmacology, Toxicology, and Pharmacy, University of Veterinary Medicine Hannover, Hannover, Germany.
  • 2 Department of Trauma Surgery, Hannover Medical School, Hannover, Germany.
  • 3 Leibniz-Institute for Molecular Pharmacology, FMP, Berlin, Germany.
  • 4 Center for Systems Neuroscience, Hannover, Germany. wolfgang.loescher@tiho-hannover.de.

摘要


BACKGROUND:Predictive in vitro models of the human blood-brain barrier (BBB) are essential in early drug discovery and development. Among available immortalized human brain capillary endothelial cell lines (BCECs), the hCMEC/D3 cell line has become the most widely used in vitro BBB model. However, monolayers of hCMEC/D3 cells form only moderately restrictive barriers, most likely because the major tight junction protein, claudin-5, is markedly downregulated. Thus, hCMEC/D3 monolayers cannot be used for vectorial drug transport experiments, which is a major disadvantage of this model. METHODS:Here we transduced hCMEC/D3 cells with a claudin-5 plasmid and compared the characteristics of these cells with those of hCMEC/D3 wildtype cells and primary cultured porcine BCECs. RESULTS:The claudin-5 transduced hCMEC/D3 exhibited expression levels (and junctional localization) of claudin-5 similar to those of primary cultured porcine BCECs. The transduced cells exhibited increased TEER values (211 Ω cm2) and reduced paracellular mannitol permeability (8.06%/h), indicating improved BBB properties; however, the barrier properties of porcine BCECs (TEER 1650 Ω cm2; mannitol permeability 3.95%/h) were not reached. Hence, vectorial transport of a selective P-glycoprotein substrate (N-desmethyl-loperamide) was not observed in claudin-5 transduced hCMEC/D3 (or wildtype) cells, whereas such drug transport occurred in porcine BCECs. CONCLUSIONS:The claudin-5 transduced hCMEC/D3 cells provide a tool to studying the contribution of claudin-5 to barrier tightness and how this can be further enhanced by additional transfections or other manipulations of this widely used in vitro model of the BBB.

KEYWORDS: P-glycoprotein, Porcine brain endothelial cells, Primary culture, Transwell