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. 2020 Aug 13;10:13708. doi: 10.1038/s41598-020-70126-7

Figure 5 .

Figure 5 

ATG7 knockdown protects against thrombin-induced EC permeability by inhibiting disassembly of VE-Cadherin at AJs. (A) HPAEC were transfected with si-Con or si-ATG7 for 24 h. Cells were then seeded on inserts placed in a 24-well plate and allowed to grow to confluence. The cells were treated with thrombin (2.5 U/ml) for 15 min and FITC-Dextran was added directly to the culture media. FITC-Dextran which had passed through the monolayer was measured using a fluorescent plate reader. Error bars represent mean ± S.E. (n = 5 for each condition). (B) HPAEC were transfected for 48 h with si-Con or si-ATG7. Cells were treated with thrombin (5 U/ml) for 15 min and non-permeabilized cells were stained with anti-VE-cadherin (green) to mark cell surface VE-cadherin and DAPI (blue) to mark nuclei. Images are representative of three experiments. (C) The percent of cells with disrupted adherens junctions (AJs) in (B) was counted as described in the “Materials and Methods” section. Error bars represent mean ± S.E. (n = 3 with 40–50 cells were counted in every fields for each conditions). (D) The percent gap area in (B) was calculated as described in the “Materials and Methods”. Error bars represent mean ± S.E. (n = 3 fields analyzed per condition).