Mycotoxin Effects on MAC-T Cell Barrier

An insert-based MAC-T cell culture system was established as previously described with slight modifications [50 (link)] to assess the paracellular permeability of MAC-T cell monolayer. MAC-T cells were seeded at 2.5 × 10⁴ cells per Transwell® insert (Corning® Transwell® #3470, 6.5 mm, 0.4 µm pore size) pre-coated with Type I collagen at 10 μg/cm² (C3867, Sigma-Aldrich) according to the manufacturer’s instruction and grown for 33 days in the same medium described above to ensure stable transepithelial electrical resistance (TEER) readings were yield according to our preliminary studies (data not shown). Medium was refreshed for both apical (AP) and basal compartments (BL) of the Transwell® inserts every other day [82 (link)]. On day 33, the cells were exposed to the increasing non-cytotoxic concentrations of either OTA (0, 2, 4.8 and 9.6 μmol/L), or CIT (0, 30, 60 and 80 μmol/L) in the AP compartment for 48 h. The TEER readings were measured before the addition of mycotoxins (TEER₀) and after 48 h mycotoxin exposure (TEER₄₈) using a Millicell ERS-2 Voltohmmeter (EMD Millipore Corporation) according to the manufacturer’s instruction. The change in TEER was expressed as TEER₄₈ to TEER₀ ratio, which was calculated according to the following formula [112 (link)]: