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. 2021 Nov 25;23(1):79. doi: 10.3892/etm.2021.11002

Figure 1.

Figure 1

Analysis of the passage of B355227 through an in vitro BBB model. (A) Confirmation of barrier establishment using TEER. bEND.3 cell cultured in Transwell inserts demonstrated a gradual and significant rise in TEER over several days in culture. *P<0.05 vs. 8 day group. (B) Cells treated with different concentrations of B355227 following the formation of the barrier revealed no adverse effects of B355227 at concentrations up to 20 µM. *P<0.05 vs. control group. (C) HPLC chromatograms demonstrate the detection of caffeine and B355227 following the permeability assay. (D) B355227 permeability assay revealed a time-dependent increase in the passage of B355227 as measured using HPLC. *P<0.05 vs. 1 h group. (E) Immunofluorescent staining of the tight junction protein ZO-1 confirms integrity of the barrier following the permeability assay. Continuous expression of ZO-1 (green) was observed along the cytoplasmic border of cells and central nuclei (blue) following implementation of the BBB permeability assay. Magnification, x40. Data are presented as the mean (n=3) ± standard deviation, obtained from three independent experiments. BBB, blood-brain barrier; TEER, trans-endothelial electrical resistance; HPLC, high-performance liquid chromatography; ZO-1, zonula occludens-1.