Isolation and Characterization of Duck Aortic Endothelial Cells

Primary duck aortic endothelial cells were isolated and characterized as previously described (25 (link)) and maintained in microvascular endothelial cell growth medium-2 (EGM-2MV; Lonza) on plates coated with 0.2% gelatin (Sigma-Aldrich) at 40°C in 5% CO₂. Furin-deficient LoVo cells (CCL-229; ATCC) were maintained in Kaighn's modification of Ham's F-12 medium (Thermo Fisher Scientific) supplemented with 10% fetal calf serum (FCS; Sigma), 100 U/mL penicillin (Lonza), and 100 μg/mL streptomycin (Lonza), at 37°C in 5% CO₂. Madin-Darby canine kidney cells (MDCK) were cultured in Eagle’s minimal essential medium (Lonza) supplemented with 10% FCS, 100 U/mL penicillin, 100 U/mL streptomycin, 2 mM l-glutamine (Lonza), 1.5 mg/mL sodium bicarbonate (Lonza), 20 mM HEPES (Lonza), and nonessential amino acids (Lonza) at 37°C in 5% CO₂. Duck lung homogenate (dLH) was obtained by digesting freshly isolated lungs of 21-day-old domestic duck embryonated eggs with collagenase and dispase (Roche) followed by treatment with a red blood cell lysis buffer (Roche) for 10 min at room temperature.