oa Sufficient Tissue Engineered Heart Valves: A Question of Cell Source?

A promising approach to solve the problems of currently used heart valve prostheses, e.g. degeneration, need for anticoagulation and risk of endocarditis, is the tissue engineering of heart valves. By using patient derived cells and fibrin as a scaffold for these valves we aim at completely autologous heart valves which have the potential to grow and hence are especially interesting for valve replacement in paediatric surgery. However, a major obstacle on the way to clinical application of tissue engineered heart valves (TEHV) is the cell-mediated tissue contraction which leads to the shrinkage of the valve’s leaflets and thus to its insufficiency. Several groups tested TEHV in the pulmonary position in the sheep model and reported mild to moderate regurgitation already at a short postimplantation time. Our goal was to analyse the influence of the cell source on the sufficiency of TEHV. Different cell sources, among which ovine carotid artery (OCA) and umbilical artery (OUA), were compared on their contractility and contraction of fibrin gels in which the cells were embedded. Cell phenotype was characterized by immunostaining of α-smooth muscle actin (α-SMA) and myosin light-chain kinase (MLCK) as markers for cell contractile activity. For the gel contraction assay, fibrin gels with a cell concentration of 5 × 10^6/ml were moulded in a 24-well plate (n ≥ 3) and their retraction was evaluated over 15 days by measuring the gels' area in relation to their original area. Hydroxyproline content, cell proliferation and burst strength were also determined. Ovine carotid artery cells exhibited a highly contractile myofibroblast phenotype (high α-SMA and MLCK expression), while OUA cells were mostly non-contractile fibroblasts with only few cells expressing α-SMA and MLCK. After 15 days, OCA embedded gels were contracted to 30.6 ± 2.0% of the original size while OUA gels maintained a size of 83.2 ± 3.7% of the initial area. To directly correlate cell contractility and valve sufficiency we moulded fibrin based heart valves using OUA and OCA cells. All valves were conditioned statically for 14 days in the closed-leaflet configuration and successively dynamically in the open-leaflet configuration in bioreactors for 30 days. While the leaflets of TEHV with OCA cells contracted and led to insufficient valves at the end of the conditioning protocol, we were able to produce completely sufficient heart valves after in vitro conditioning using non-contractile OUA cells. In an on-going animal study OUA embedded TEHV, after being seeded with endothelial cells, are implanted in the pulmonary artery in lambs to analyse their growth potential and their hemodynamic performance in vivo. Transesophageal echocardiography showed both in colour Doppler and in cw Doppler mode that after fourteen weeks a first implanted valve was still completely sufficient and showed no sign of cell-mediated tissue contraction.