| We sought to minimize the requirements for maintaining the well-being and pluripotence of human embryonic stem cell (hESC). We identified bFGF (at an optimal concentration of 20 ng/ml), insulin (at a supraphysiological concentration of 20 mug/ml), ascorbic acid, and laminin as the minimal essential components of a defined culture system that could maintain clonal hESCs in a stable, healthy, self-renewing, pluripotent state. Providing small molecule inducers enabled our system to serve as a platform for directing such hESCs towards particular differentiated cell types efficiently and exclusively, including electrophysiologically-coordinated cardiomyocytes and neurons, without an intervening multi-lineage embryoid body (EB) stage. Neural progenitors derived in this fashion were safely engraftable. It appears that this formulation permits the spontaneous unfolding of inherent embryogenesis-like processes that, in turn, aid efficient clonal expansion and de novo derivation of stable hESCs from blastocysts. |
