Three-dimensional co-culture facilitates the differentiation of embryonic stem cells into mature cardiomyocytes

✍ Scribed by Dong-Bo Ou; Yong He; Rui Chen; Ji-Wei Teng; Hong-Tao Wang; Di Zeng; Xiong-Tao Liu; Lu Ding; Jin-Yan Huang; Qiang-Sun Zheng

• Publisher: John Wiley and Sons
• Year: 2011
• Tongue: English
• Weight: 563 KB
• Volume: 112
• Category: Article
• ISSN: 0730-2312
• DOI: 10.1002/jcb.23283

✦ Synopsis

The cardiomyocyte (CM) differentiation of embryonic stem cells (ESCs) is routinely cultured as two-dimensional (2D) monolayer, which doesn't mimic in vivo physiological environment and may lead to low differentiated level of ESCs. Here, we develop a novel strategy that enhances CM differentiation of ESCs in collagen matrix three-dimensional (3D) culture combined with indirect cardiac fibroblasts co-culture. ESCs were cultured in hanging drops to form embryoid bodies (EBs) and then applied on collagen matrix. The EBs were indirectly co-cultured with cardiac fibroblasts by the hanging cell culture inserts (PET 1 mm). The molecular expressions and ultrastructural characteristics of ESCderived CMs (ESCMs) were analyzed by real time RT-PCR, immunocytochemistry, and Transmission Electron Microscopy (TEM). We found that the percentage of beating EBs with cardiac fibroblasts co-culture was significantly higher than that without co-culture after differentiation period of 8 days. Type I collagen used as 3D substrates enhanced the late-stage CM differentiation of ESCs and had effect on ultrastructural mature of ESCMs in late-stage development. The combined effects of 3D and co-culture that mimic in vivo physiological environment further improved the efficiency of CM differentiation from ESCs, resulting in fiber-like structures of cardiac cells with organized sarcomeric structure in ESCMs. This novel 3D co-culture system emphasizes the fact that the ESC differentiation is actively responding to cues from their environment and those cues can drive phenotypic control, which provides a useful in vitro model to investigate CM differentiation of stem cells.