Directed differentiation of mouse embryonic stem cells to cardiomyocytes in a scalable culture system.

摘要

Heart disease is top ranked as cause of morbidity and mortality in the US and most developed countries. Myocardial infarction is associated with significant cardiomyocyte death and permanently impaired cardiac function. Widespread utility of heart transplantation is hindered due to severe shortage of donor organs. Hence, the search for alternative sources of heart cells including embryonic stem cells (ESCs) has intensified in recent years.;We have explored methodologies for the cardiogenic differentiation of ESCs. We have examined traditional differentiation techniques in detail. We discuss their limitations, present optimization strategies and explore novel approaches with potential to surpass these limitations. Specifically, we explored the differentiation of ESCs towards cardiomyocytes in the absence of serum and with factors involved in embryonic heart development. Current methods for ESC-to-cardiomyocyte differentiation rely on the use of serum which makes challenging the control of ESC specification, for example, via the addition of physiologically relevant agents. The resulting cell populations are heterogeneous and contain only minute fractions of cells displaying cardiomyocyte markers. We have identified conditions using defined serum-free medium and TGF-beta ligands (mainly bone morphogenetic proteins; BMP) for directing the differentiation of mouse ESCs (mESCs) to cardiomyocyte-like cells. The BMP directed differentiation offers advantages over the traditional techniques such as reproducible results and higher beating percentages. Given the need for generating heart cells in adequate quantities for clinical uses a method was subsequently developed for directing the differentiation of mESCs in a stirred-suspension bioreactor without serum.;These findings will be translated to human ESC (hESC) differentiation aiming at the development of a scalable system for the generation of cardiomyocytes for cell therapies. These studies also encourage exploration of other factors under defined serum-free conditions to design strategies for obtaining higher yield and reproducible differentiation of ESCs to cardiomyocytes.