Collagenase

Hepes, methionine (Met), phenylmethanesulfonyl fluoride, and 5-formyltetrahydrofolate (folinic acid (FnA)) were obtained from Sigma-Aldrich (St Louis, MO, USA). l-glutamine was purchased from Biochrom-AG (Berlin, Germany). Newborn calf bovine serum and endothelial cell growth factor were from Roche (Mannheim, Germany) and collagenase, M199 basal culture medium (with Earle’s balanced salt solution and Hepes), and Hank’s balanced salt solution were from Promocell (Heidelberg, Germany).

The human umbilical cord was collected in a transport buffer solution beaker and kept at 4 °C for 4 h. The umbilical cord was cleaned properly with PBS, and the remaining blood clots were removed. To harvest the smooth muscle cells, the arteries were isolated from the cord and minced into small ring-shaped pieces with a scalpel. The small pieces were distributed horizontally inside a T75 cell culture flask and supplied with a fresh DMEM medium (Thermofischer Scientific).
For the endothelial cells, the lumen of the HU artery was soaked with collagenase (Thermofischer Scientific) for 30 min at 37 °C for incubation. After the collagenase removal, the cells were cultured in the flask and supplied with an endothelial basal medium (Promocell, Heidelberg, Germany). The umbilical cord was collected from the Gynecology Department at the University Hospital Aachen in accordance with the human subjects’ approval of the ethics committee (vote of the local ethics committee: ‘EK 2067).