Method to direct differentiation of pluripotent stem cells into functional heart muscle

A method for producing bioengineered heart muscle from pluripotent stem cells, comprising the steps of

• (i) cultivating pluripotent stem cells in a basal medium comprising an effective amount of (a) BMP4, Activin A, FGF2, a GSK3-inhibitor, and (b) a serum-free supplement resulting in a final concentration of 0.5-50 mg/ml albumin, 1-100 μg/ml transferrin, 0.1-10 μg/ml ethanol amine, 0.003-0.3 μg/ml sodium selenite, 0.4-40 μg/ml L-Carnitine HCl, 0.1-10 μg/ml Hydrocortisone, 0.05-5 μl/ml Fatty acid supplement, and 0.0001-0.1 μg/ml triodo-L-thyronine (T3), thereby inducing mesoderm differentiation of said pluripotent stem cells;
• (ii) cultivating the cells obtained in step (i) in a basal medium comprising an effective amount of an inhibitor of the Wnt-signaling pathway and a serum-free supplement as in (i), thereby inducing cardiac differentiation of the cells;
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• (iii) cultivating the cells obtained in step (ii) in a basal medium comprising an effective amount of a serum-free supplement as in (i), under mechanical stimulation, thereby promoting cardiac maturation, wherein prior to step (i) said pluripotent stem cells are seeded in the presence of callogen in a mould.