Collagenase type 2

We collected human umbilical cords from consented de-identified patients through IRB-approved tissue procurement facility (BioNet) at the University of Minnesota. Umbilical cords were rinsed twice with Hank’s buffered salt solution and one end of the cord was sealed and the umbilical veins were filled with 10 ml of collagenase type II (1 U/ml, STEM CELL Technologies Cambridge USA). The digestion was done at 37 °C for 15 minutes in the incubator. Detached cells were then released by flushing the veins with HBSS. The mixture was centrifuged at 1500 rpm for 10 minutes and the cell pellets were resuspended in medium Endothelial Cell Growth Medium MV2 (PromoCell, USA). The medium was replaced every 48 hours. Cells were sub-cultured after treatment with 0.01% trypsin/EDTA.

Sodium chromate, glutamine (for Seahorse tests), oligomycin, 2-deoxyglucose, rotenone, and antimycin A were from Sigma-Aldrich (St. Louis, MO, USA). Trypsin/EDTA, sodium pyruvate, penicillin/streptomycin, LHC media, and L-glutamine were from Thermo Fisher Scientific (Waltham, MA, USA). Dulbecco’s minimal essential medium and Ham’s F-12 medium (DMEM/F-12) were from Mediatech (Herndon, VA, USA). Cosmic calf serum was from Hyclone (Logan, UT, USA). Collagenase Type II was from STEMCELL Technologies (Vancouver, Canada) Tissue culture dishes, flasks, and plastic ware were from Falcon Labware (Becton-Dickinson, Franklin Lakes, NJ, USA). Carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP) and XF base medium were from Agilent Technologies (Santa Clara, CA, USA).