Figure 7.

Permeabilities of paracellular, macromolecular, and transcytotic flux markers. Permeability measurements of paracellular markers revealed a passage of macromolecules in a range from ∼400–900 Da up to 10–20 kDa in tricellular overexpression clones, whereas bicellular overexpression clones showed permeation already for smaller molecules. (A) Fluorescein (332 Da) was used as a twice negative–charged paracellular flux marker. In comparison with the vector-transfected cells permeabilities were decreased in TRa-8 cells and remained unchanged in TRa-4 cells (n = 6, 9, and 10; ** p < 0.01). (B) Permeability for PEG-400 remained similar in mock-transfected and TRa-4 cells, whereas it was decreased in TRa-8 cells (n = 9, 12, and 11; ** p < 0.01). (C) PEG-900 permeabilities for TRa-8 and also TRa-4 cells were reduced (n = 4 and 6; * p < 0.05; and n = 7; ** p < 0.01, respectively). (D) Permeability for FITC-dextran 4 kDa (FD4) was strongly decreased already in TRa-4 cells and also in TRa-8 cells (n = 6 and 6; ** p < 0.01; and n = 8; *** p < 0.001). (E) Permeability for FITC-dextran 10 kDa (FD10) showed a pattern similar to that of FD4, with even lower values in TRa-8 cells (n = 5 each; ** p < 0.01). (F) Permeability for FD20 (10⁻⁹ cm/s) in control cells was 20 times lower than that for FD10 and was not significantly altered in both tricellulin-transfected clones (n = 6, 5, and 6). (G) Permeability for the transcytotic marker HRP. Permeabilities for HRP (10⁻¹² cm/s) remained the same in mock- and both tricellulin-transfected clones, indicating that the rate of transcytosis is not changed because of TRIC-a overexpression (n = 4, 5, and 4). (H) An unchanged percentage of TUNEL-positive cells revealed no changes in apoptosis because of overexpression of TRIC-a (n = 6).