No abstract

We describe a new, efficient protocol that involves the serial addition of noggin, basic fibroblast growth factor (bFGF), retinoic acid, and sonic hedgehog (Shh) for the differentiation of human induced pluripotent stem cells (hiPSC) to retinal pigmented epithelium (RPE) in a serum- and feeder-free adherent condition. hiPSC-RPE cells exhibited RPE morphology and specific molecular markers. Additionally, several hiPSC lines were generated from retinal-specific patients with Leber's congenital amaurosis, Usher syndrome, two patients with retinitis pigmentosa, and a patient with Leber's hereditary optic neuropathy. The RPE cells generated from these disease-specific hiPSCs expressed specific markers by the same RPE lineage-directed differentiation protocol. These findings indicate a new short-term, simple, and efficient protocol for differentiation of hiPSCs to RPE cells. Such specific retinal disease-specific hiPSCs offer an unprecedented opportunity to recapitulate normal and pathologic formation of human retinal cells in vitro, thereby enabling pharmaceutical screening, and potentially autologous cell replacement therapies for retinal diseases.

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Stem cell technology combined with nano-scaffold surfaces provides a new tool for better induction involved in cell lineage differentiations and therefore for central nervous system repair. This study was undertaken to investigate appropriate neural cell-substrate interactions. Neural progenitors (NPs) were established from human embryonic stem cells (hESCs), as a first step, using an adherent system and a defined medium supplemented with a combination of factors. Next, the behavior of hESC-derived NPs (hESC-NPs) was evaluated on a synthetic, randomly oriented, three-dimensional nanofibrillar matrix composed of electrospun polyamide nanofibers (Ultra-Web™) using a variety of experimental approaches. We have demonstrated that homogenous, expandable, and self-renewable NPs can be easily generated from hESCs; they can express related markers Nestin, Sox1, and Pax6; and they can undergo multipotency differentiation to neurons and glials. Functionally, NPs cultured on nanofibers demonstrated an increase in the rate of migration, proliferation, morphology, and neurite length when compared with NPs cultured on two-dimensional culture surfaces. The results suggest that topographical features of the extracellular matrix of the cell environment have paved the way for a better understanding of human neuronal development, thus allowing for future clinical applications.