Supplementary Material for: Tissue Engineering of Vein Valves Based on Decellularized Natural Matrices

Valvular repair or transplantation, designed to restore the venous valve function of the legs, has been proposed as treatment in chronic venous insufficiency. Available grafts or surgeries have provided limited durability so far. Generating venous valve substitutes by means of tissue engineering could be a solution. We generated decellularized jugular ovine vein conduits containing valves (oVVC) after reseeding with ovine endothelial cells differentiated from peripheral blood-derived endothelial cells (oPBEC), cultivated in vitro corresponding to the circulatory situation in the lower leg at rest and under exertion. oVVC were decellularized by detergent treatment. GFP-labeled oPBEC were seeded onto the luminal side of the decellularized oVVC and cultivated under static-rotational conditions for 6 h (group I) and 12 h (group II), respectively. Reseeded matrices of group I were exposed to continuous low flow conditions (“leg at rest”). The tissues of group II were exposed to a gradually increasing flow (“leg under effort”). After 5 days, the grafts of group I revealed a uniform luminal endothelial cell coverage of the examined areas of the venous walls and adjacent venous valve leaflets. In group II, the cell coverage on luminal areas of the venous wall parts was found to be nearly complete. The endothelial cell coverage of adjacent venous valve leaflets was revealed to be less dense and confluent. Endothelial cells cultured on acellular vein tissues of both groups were distinctly orientated uniformly in the flow direction, clearly creating a stable and flow-orientated layer. Thus, an endothelium could successfully be reestablished on the luminal surface of a decellularized venous valve by seeding peripheral blood endothelial cells and culturing under different conditions.