Technologies, Methods, and Products of Small Molecule Directed Tissue and Organ Regeneration from Human Pluripotent Stem Cells

摘要

Pluripotent human embryonic stem cells (hESCs) hold great potential for restoring tissue and organ function, which has been hindered by inefficiency and instability of generating desired cell types through multi-lineage differentiation. This instant invention is based on the discovery that pluripotent hESCs maintained under defined culture conditions can be uniformly converted into a specific lineage by small molecule induction. Retinoic acid induces specification of neuroectoderm direct from the pluripotent state of hESCs and triggers progression to neuronal progenitors and neurons efficiently. Similarly, nicotinamide induces specification of cardiomesoderm direct from the pluripotent state of hESCs and triggers progression to cardiac precursors and cardiomyocytes efficiently. This technology provides a large supply of clinically-suitable human neuronal or cardiac therapeutic products for CNS or myocardium repair. This invention enables well-controlled efficient induction of pluripotent hESCs exclusively to a specific clinically-relevant lineage for tissue and organ engineering and regeneration, cell-based therapy, and drug discovery.

主权项

1. A method of producing cells of neuronal lineage directly from pluripotent human stem cells, comprising steps of:
(i) providing a culture of pluripotent human stem cells in a defined medium comprising bFGF, insulin, ascorbic acid, and activin-A, wherein said defined medium is free of serum, free of feeder cells, and free of feeder cell conditioned medium; (ii) adding retinoic acid (RA) to the culture, wherein the pluripotent human stem cells are cultured in the presence of RA for a period of time sufficient to cause Nurr-1 to translocate to the nucleus and cause the pluripotent human stem cells to directly differentiate into cells of neuronal lineage comprising at least 90% of neuroectodermal cells; (iii) further differentiating the neuroectodermal cells into a population of cells comprising at least 90% of neuronal progenitor cells; and (iv) further differentiating the neuronal progenitor cells into a population of cells comprising at least 90% of neuronal cells.