UNIT 1D.7 Differentiation of Human Embryonic Stem Cells to Cardiomyocytes on Microcarrier Cultures

Abstract

We have developed an improved cardiomyocyte differentiation protocol where we stabilized embryoid bodies (EB) in serum- and insulin-free medium (bSFS) supplemented with p38 MAP kinase inhibitor (SB203580) by addition of 10 µm laminin-coated positively charged (protamine sulfate derivatized TSKgel Tresyl-5PW) microcarriers. This protocol achieved a maximum 3-fold cell expansion, differentiation efficiency of 20%, and an overall cardiomyocyte yield of 3 × 10⁵ CM/ml in static conditions. In comparison, EB cultures achieved 1.5-fold cell expansion, differentiation efficiency of 15%, and an overall cardiomyocyte yield of 1.1 × 10⁵ CM/ml. The scalability of this platform was demonstrated in suspended spinner cultures, producing a maximum of 2.14 × 10⁵ CM/ml in 50-ml cultures. This yield is two-fold higher than the control static EB-based platform (1.1 × 10⁵ CM/ml), and seven-fold higher than yields reported in literature, 3.1–9 × 10⁴ CM/ml. The robustness of this protocol was tested with HES-3 and H1 cell lines. Curr. Protoc. Stem Cell Biol. 21:1D.7.1-1D.7.14. © 2012 by John Wiley & Sons, Inc.