Endohm ohmmeter

To establish polarized pBCEC cultures (Kober et al., 2017 (link)), cells were trypsinized and plated onto collagen‐coated (120 μg ml⁻¹) 12‐well transwell filter plates (0.4‐μm pore size, Corning) at a density of 40,000 cells cm^‐2. Cells were grown for 2–3 days depending on the transendothelial electrical resistance (TEER; 50 Ω cm⁻²). The tightness of the transwell culture was assessed by measuring TEER using an EndOhm tissue resistance measurement chamber and EndOhm ohmmeter (World Precision Instruments, Florida). TEER of collagen‐coated, cell‐free filters were used as blanks. Tight junction formation was induced (overnight) by adding DMEM/Ham's F‐12 medium containing 550 nM hydrocortisone, 1% penicillin/streptomycin and 0.7 mM l‐glutamine. Establishment of intact tight junctions was indicated by TEER rising above 150 Ω cm⁻². Agents known to increase BBB permeability, 27OH (0.055 and 0.5 μM), 24OH (0.5 μM), 7β‐hydroxycholesterol (7βOH; 0.5 μM) or LPS (5 μM), were used as positive controls and were added to apical media containing 1% BSA. TEER were measured at indicated time points for up to 24 h. Six independent experiments with the above additions and use of aliquots from the same cell culture were performed (Figure 5). In addition, two separate independent experiments with other cell cultures were performed with n = 3.