HUVECs were thawed and resuspended in activation mix, and seeded in 96-well tissue culture plates (1 × 104 cells/well). After 6 h, fresh control medium was added to “starve” the cells for 18–20 h. The day after, the medium was replaced with fresh control medium, activation mix (positive control), or control medium supplemented with recombinant (r) human VEGFA (250 ng/mL, PeproTech Nordic), or diluted mucinous tumor tissue from PMP patients and then incubated for 72 h. Determination of cell viability was performed using the MTS assay as previously described [26]. The HUVEC proliferation assay results were calculated from six replicates in each experiment with three to eleven independent experiments (Figure 2a). To assess the effect of adding anti-angiogenic drugs, tissue samples were pre-incubated with BEV (0.25 or 2.5 mg/mL) at 37 °C for 2 h before transfer to the HUVEC cells. HUVECs were then incubated, as described above. The experiment was repeated at least 4 times.

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